U.S. patent number 5,929,040 [Application Number 08/666,392] was granted by the patent office on 1999-07-27 for method for the prophylaxis and/or treatment of proliferative and/or inflammatory skin disorders.
This patent grant is currently assigned to Royal Children's Hospital Research Foundation. Invention is credited to George Arthur Werther, Christopher John Wraight.
United States Patent |
5,929,040 |
Werther , et al. |
July 27, 1999 |
Method for the prophylaxis and/or treatment of proliferative and/or
inflammatory skin disorders
Abstract
The present invention relates generally to a method for the
prophylaxis and/or treatment of skin disorders, and in particular
proliferative and/or inflammatory skin disorders, and to genetic
molecules useful for same. The present invention is particularly
directed to genetic molecules capable of modulating growth factor
interaction with its receptor on epidermal keratinocytes to
inhibit, reduce or otherwise decrease stimulation of this layer of
cells. The present invention contemplates, in a most preferred
embodiment, a method for the prophylaxis and/or treatment of
psoriasis.
Inventors: |
Werther; George Arthur
(Camberwell, AU), Wraight; Christopher John
(Blackburn, AU) |
Assignee: |
Royal Children's Hospital Research
Foundation (Victoria, AU)
|
Family
ID: |
3781291 |
Appl.
No.: |
08/666,392 |
Filed: |
August 20, 1996 |
PCT
Filed: |
July 06, 1995 |
PCT No.: |
PCT/AU95/00410 |
371
Date: |
August 20, 1996 |
102(e)
Date: |
August 20, 1996 |
PCT
Pub. No.: |
WO96/01636 |
PCT
Pub. Date: |
January 25, 1996 |
Foreign Application Priority Data
Current U.S.
Class: |
514/44A |
Current CPC
Class: |
A61K
31/7088 (20130101); A61P 17/06 (20180101); A61P
43/00 (20180101); A61K 31/7105 (20130101); A61K
31/7125 (20130101); A61K 31/711 (20130101); A61P
17/00 (20180101); A61P 35/00 (20180101); A61P
29/00 (20180101) |
Current International
Class: |
A61P
17/06 (20060101); A61P 17/00 (20060101); C07K
14/475 (20060101); A61K 31/7088 (20060101); A61K
31/70 (20060101); A61K 38/18 (20060101); A61K
9/10 (20060101); C12N 15/09 (20060101); A61K
9/06 (20060101); A61K 48/00 (20060101); C12N
9/22 (20060101); C07H 21/00 (20060101); C12N
15/11 (20060101); C07H 21/04 (20060101); C07H
21/02 (20060101); C07K 14/435 (20060101); A61K
031/70 () |
Field of
Search: |
;435/6,91.31,325,375
;514/44 ;536/24.31,24.5 |
References Cited
[Referenced By]
U.S. Patent Documents
Foreign Patent Documents
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|
|
|
|
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|
9422486 |
|
Oct 1994 |
|
WO |
|
9423034 |
|
Oct 1994 |
|
WO |
|
9610401 |
|
Apr 1996 |
|
WO |
|
Other References
Antisense '97: A roundtable on the state of the industry. Nature
Biotechnology 15: 519-524, Jun. 1997. .
Gewirtz et al. Facilitating oligonucleotide delivery: Helping
antisense deliver on its promise. Proc. Natl. Acad. Sci. USA 93:
3161-3163, Apr. 1996. .
Gura. Antisense has growing pains. Science 270: 575-577, Oct. 1995.
.
Stull et al. Antigene, ribozyme and aptamer nucleic acid drugs:
Progress and prospects. Pharm. Res. 12: 465-483, Apr. 1995. .
Rojanasakul. Antisense oligonucleotide therapeutics: Drug delivery
and targeting. Adv. Drug Del. Rev. 18:115-131, 1996. .
Gouy, M. Secondary structure prediction of RNA, in Nucleic Acid and
Protein Sequence Analysis: A Practical Approach, Bishop et al, eds.
IRL Press, Oxford, pp. 259-284, 1987. .
"Localization of Messenger Ribonucleic Acid for Insulin-Like Growth
Factor-Binding Proteins in Human Skin by in Situ Hybridization" by
J.A. Batch et al.; Journal of Clinical Endocrinology and
Metabolism; vol. 79, No. 5, pp. 1444-1449. 1994. .
"Regulation of IGFBP Secretion and Modulation of Cell Growth in
MDBK Cells" by W.S. Cohick et al.; pp. 20-23; University of North
Carolina School of Medicine, Division of Endocrinology. 1993. .
"Episomal Expression of Sense and Antisense Insulin-like Growth
Factor (IGF)-binding Protein-4 Complementary DNA Alters the
Mitogenic Response of a Human Colon Cancer Cell Line (HT-29) by
Mechanisms that are Independent of an Dependent upon IGF-1" by
Singh et al.; Cancer Research 54, pp. 6563-6570, Dec. 15, 1994.
.
"Loss of the Metastatic Phenotype in Murine Carcinoma Cells
Expressing an Antisense RNA to the Insulin-like Growth Factor
Receptor" by Long et al.; Cancer Research 55, pp. 1006-1009, Mar.
1, 1995. .
"Growth Inhibition of Human Melanoma Cells in Nude Mice by an
Antisense Strategies to the Type 1 Insulin-like Growth Factor
Receptor" by Resnicoff et al.; Cancer Research 54, pp. 4848-4850,
Sep. 15, 1994. .
"Antisense-Mediated Reduction in Insulin-Like Growth Factor-1
Receptor Expression Suppresses the Malignant Phenotype of a Human
Alveolar Rhabdomyosarcoma" by Shapiro et al.; The American Society
for Clinical Investigation, Inc.; vol. 94, Sep. 1994, pp.
1235-1242. .
"The Insulin-like Growth Factor 1 Receptor Is Expressed by
Epithelial Cells with Proliferative Potential in Human Epidermis
and Skin Appendages: Correlation of Increased Expression with
Epidermal Hyperplasia" by Hodak et al.; Laboratory for
Investigative Dermatology; pp. 564-570. 1996..
|
Primary Examiner: Degen; Nancy
Assistant Examiner: Larson; Thomas G.
Attorney, Agent or Firm: Baker & Botts
Parent Case Text
This is a national stage filing of PCT/AU95/00410, filed Jul. 6,
1995.
Claims
We claim:
1. A method for ameliorating the effects of an IGF-I mediated
proliferative, inflammatory, or proliferative and inflammatory
disorder in the skin of a mammal, said method comprising contacting
the skin of a mammal in need of such treatment with a nucleic acid
molecule or chemical analog thereof that hybridizes to a nucleic
acid encoding the IGF-I receptor and thereby inhibits expression of
the IGF-I receptor, in an amount effective to reduce proliferation,
inflammation, or proliferation and inflammation of the skin of said
mammal.
2. The method of claim 1 wherein said nucleic acid encoding the
IGF-I receptor has the sequence of SEQ ID NO:3.
3. The method of claim 1 wherein said disorder is psoriasis.
4. The method of claim 1 wherein said mammal is a human.
5. The method of claim 1 wherein said nucleic acid molecule is
about fifteen nucleotides in length.
6. A method for ameliorating the effects of an IGF-I mediated
proliferative, inflammatory, or proliferative and inflammatory
disorder in the skin of a mammal, said method comprising contacting
the skin of a mammal in need of such treatment with a nucleic acid
molecule or chemical analog thereof that hybridizes to a nucleic
acid encoding IGFBP-3 and thereby inhibits expression of IGFBP-3,
in an amount effective to reduce proliferation, inflammation, or
proliferation and inflammation of the skin of said mammal.
7. The method of claim 6 wherein said nucleic acid encoding IGFBP-3
has the sequence of SEQ ID NO:2.
8. The method of claim 6 wherein said disorder is psoriasis.
9. The method of claim 6 wherein said mammal is a human.
10. The method of claim 6 wherein said nucleic acid molecule is
about fifteen nucleotides in length.
11. A method of treatment of psoriasis comprising contacting the
skin of a mammal in need of such treatment with a nucleic acid
molecule or chemical analog thereof that hybridizes to a nucleic
acid encoding the IGF-I receptor and thereby inhibits expression of
the IGF-I receptor, in an amount effective to reduce cell
proliferation of the skin of said mammal.
12. The method of claim 11 wherein said nucleic acid encoding the
IGF-I receptor has the sequence of SEQ ID NO:3.
13. The method of claim 11 wherein said disorder is psoriasis.
14. The method of claim 11 wherein said mammal is a human.
15. The method of claim 11 wherein said nucleic acid molecule is
about fifteen nucleotides in length.
16. A method of treatment of psoriasis comprising contacting the
skin of a mammal in need of such treatment with a nucleic acid
molecule or chemical analog thereof that hybridizes to a nucleic
acid encoding IGFBP-3 and thereby inhibits expression of IGFBP-3,
in an amount effective to reduce cell proliferation of the skin of
said mammal.
17. The method of claim 16 wherein said nucleic acid encoding
IGFBP-3 has the sequence of SEQ ID NO:2.
18. The method of claim 16 wherein said disorder is psoriasis.
19. The method of claim 16 wherein said mammal is a human.
20. The method of claim 16 wherein said nucleic acid molecule is
about fifteen nucleotides in length.
Description
The present invention relates generally to a method for the
prophylaxis and/or treatment of skin disorders, and in particular
proliferative and/or inflammatory skin disorders, and to genetic
molecules useful for same. The present invention is particularly
directed to genetic molecules capable of modulating growth factor
interaction with its receptor on epidermal keratinocytes to
inhibit, reduce or otherwise decrease stimulation of this layer of
cells. The present invention contemplates, in a most preferred
embodiment, a method for the prophylaxis and/or treatment of
psoriasis.
Bibliographic details of the publications numerically referred to
in this specification are collected at the end of the description.
Sequence Identity Numbers (SEQ ID NOs.) for the nucleotide
sequences referred to in the specification are defined following
the bibliography.
Throughout this specification, unless the context requires
otherwise, the word "comprise", or variations such as "comprises"
or "comprising", will be understood to imply the inclusion of a
stated element or integer or group of elements or integers but not
the exclusion of any other element or integer or group of elements
or integers.
Psoriasis and other similar conditions are common and often
distressing proliferative and/or inflammatory skin disorders
affecting or having the potential to affect a significant
proportion of the population. The condition arises from over
proliferation of basal keratinocytes in the epidermal layer of the
skin associated with inflammation in the underlying dermis. Whilst
a range of treatments have been developed, none is completely
effective and free of adverse side effects. Although the underlying
cause of psoriasis remains elusive, there is some consensus of
opinion that the condition arises at least in part from over
expression of local growth factors and their interaction with their
receptors supporting keratinocyte proliferation via keratinocyte
receptors which appear to be more abundant during psoriasis.
One important group of growth factors are the dermally-derived
insulin-like growth factors (IGFs) which support keratinocyte
proliferation. In particular, IGF-I and IGF-II are ubiquitous
peptides each with potent mitogenic effects on a broad range of
cells. Molecules of the IGF type are also known as "progression
factors" promoting "competent" cells through DNA synthesis. The
IGFs act through a common receptor known as the Type I or IGF-I
receptor, which is tyrosine kinase linked. They are synthesised in
mesenchymal tissues, including the dermis, and act on adjacent
cells of mesodermal, endodermal or ectodermal origin. The
regulation of their synthesis involves growth hormone (GH) in the
liver, but is poorly defined in most tissues (1).
Particular proteins, referred to as IGF binding proteins (IGFBPs),
appear to be involved in autocrine/paracrine regulation of tissue
IGF availability (2). Six IGFBPs have so far been identified. The
exact effects of the IGFBPs is not clear and observed effects in
vitro have been inhibitory or stimulatory depending on the
experimental method employed (3). There is some evidence, however,
at certain IGFBPs are involved in targeting IGF-I to its cell
surface receptor.
Skin, comprising epidermis and underlying dermis, has GH receptors
on dermal fiboblasts (4). Fibroblasts synthesize IGF-I as well as
IGFBPs-3, -4, -5 and -6 (5) which may be involved in targeting
IGF-I to adjacent cells as well as to the overlaying epidermis. The
major epidermal cell type, the keratinocyte, does not synthesize
IGF-I, but possesses IGF-I receptors and is responsive to IGF-I
(6).
It is apparent, therefore, that IGF-I and other growth promoting
molecules, are responsible for or at least participate in a range
of skin cell activities. In accordance with the present invention,
the inventors have established that abberations in the normal
functioning of these molecules or abberations in their interaction
with their receptors is an important factor in proliferative and/or
inflammatory skin disorders. It is proposed, therefore, to target
these molecules or other molecules which facilitate their
functioning or interaction with their receptors to thereby
ameliorate the effects of abberant activity during or leading to
skin disease conditions.
Accordingly, one aspect of the present invention contemplates a
method for ameliorating the effects of a proliferative and/or
inflammatory skin disorder in a mammal, said method comprising
contacting the proliferating and/or inflamed skin or skin capable
of proliferation and/or inflammation with an effective amount of a
nucleic acid molecule or chemical analogue thereof capable of
inhibiting or otherwise reducing a growth factor mediated cell
proliferation and/or inflammation.
Growth factor mediated cell proliferation and inflammation are also
referred to as epidermal hyperplasias and may be mediated by any
number of molecules such as but not limited to IGF-I, keratinocyte
growth factor (KGF), transforming growth factor-.alpha.
(TGF.alpha.), tumour necrosis factor-.alpha. (TNF.alpha.),
interleukin-1, -4, -6 and 8 (IL-1, IL-4, IL-6 and IL-8,
respectively), basic fibroblast growth factor (bFGF) or a
combination of one or more of the above. The present invention is
particularly described and exemplified with reference to IGF-I and
its receptor (IGF-I receptor) and to IGF-I facilitating molecules,
IGFBPs, since targeting these molecules according to the methods
contemplated herein provides the best results to date. This is
done, however, with the understanding that the present invention
extends to any growth factor or cytokine-like molecule, a receptor
thereof or a facilitating molecule like the IGFBPs involved in skin
cell proliferation such as those molecules contemplated above
and/or their receptors and/or facilitating molecules therefor.
According to is preferred embodiment, there is provided a method
for ameliorating the effects of a proliferative and/or inflammatory
skin disorder in a mammal, said method comprising contacting the
proliferating and/or inflamed skin or skin capable of proliferation
and/or inflammation with an effective amount of a nucleic acid
molecule or chemical analogue thereof capable of inhibiting or
otherwise reducing IGF-I mediated cell proliferation and/or
inflammation.
The present invention is particularly described by psoriasis as the
proliferative skin disorder. However, the subject invention extends
to a range of proliferative and/or inflammatory skin disorders or
epidermal hyperplasias such as but not limited to psoriasis,
ichthyosis, pityriasis rubra pilaris ("PRP"), seborrhoea, keloids,
keratoses, neoplasias and scleroderma, warts, benign growths and
cancers of the skin.
In a preferred embodiment, therefore, the present invention is
directed to a method for ameliorating the effects of psoriasis,
said method comprising contacting proliferating skin or skin
capable of proliferation with an effective amount of a nucleic acid
molecule or chemical analogue thereof capable of inhibiting or
otherwise reducing IGF-I mediated cell proliferation.
The present invention extends to any mammal such as but not limited
to humans, livestock animals (e.g. horses, sheep, cows, goats,
pigs, donkeys), laboratory test animals (e.g. rabbits, mice, guinea
pigs), companion animals (e.g. cats, dogs) and captive wild
animals. However, the instant invention is particularly directed to
proliferative and/or inflammatory skin disorders such as psoriasis
in humans.
The aspects of the subject invention instantly contemplated are
particularly directed to the topical application of one or more
suitable nucleic molecules capable of inhibiting, reducing or
otherwise interfering with IGF-mediated cell proliferation and/or
inflammation. More particularly, the nucleic acid molecule targets
IGF-I interaction with its receptor. Conveniently, therefore, the
nucleic acid molecule is an antagonist of IGF-I interaction with
its receptor. Most conveniently, the nucleic acid molecule
antagonist is an antisense molecule to the IGF-I receptor, to IGF-I
itself or to a molecule capable of facilitating IGF-I interaction
with its receptor such as but not limited to an IGFBP.
Insofar as the invention relates to IGFBPs, the preferred molecules
are IGFBP-2, -3, -4, -5 and -6. The most preferred molecules are
IGFBP-2 and IGFBP-3.
The nucleotide sequences of IGFBP-2 and IGFBP-3 are set forth in
FIGS. 1 (SEQ ID NO. 1) and 2 (SEQ ID NO. 2), respectively.
According to a particularly preferred aspect of the present
invention, there is provided a nucleic acid molecule comprising at
least about ten nucleotides capable of hybridising to, forming a
heterodouplex or otherwise interacting with an mRNA molecule
directed from a gene corresponding to a genomic form of SEQ ID NO.
1 and/or SEQ ID NO. 2 and which thereby reduces or inhibits
translation of said mRNA molecule. Preferably, the nucleic acid
molecule is at least about 15 nucleotides in length and more
preferably at least about 20-25 nucleotides in length. However, the
instant invention extends to any length nucleic acid molecule
including a molecule of 100-200 nucleotides in length to correspond
to the full length of or near full length of the subject genes.
The nucleotide sequence of the antisense molecules may correspond
exactly to a region or portion of SEQ ID NO. 1 or SEQ ID NO. 2 or
may differ by one or more nucleotide substitutions, deletions
and/or additions. It is a requirement, however, that the nucleic
acid molecule interact with an mRNA molecule to thereby reduce its
translation into active protein.
Examples of potential antisense molecules for IGFBP-2 and IGFBP-3
are those capable of interacting with sequences selected from the
lists in Examples 6 and 7, respectively.
The nucleic acid molecules in the form of an antisense molecule may
be linear or covalently closed circular and single stranded or
partially double stranded. A double stranded molecule may form a
triplex with target mRNA or a target gene. The molecule may also be
protected from, for example, nucleases, by any number of means such
as using a nonionic backbone or a phosphorothioate linkage. A
convenient nonionic backbone contemplated herein is
ethylphosphotriester linkage or a 2'-O-methylribosyl
derivative.
Examples of suitable oligonucleotide analogues are conveniently
described in Ts'O et al (7).
Alternatively, the antisense molecules of the present invention may
target the IGF-I gene itself or its receptor or a multivalent
antisense molecule may be constructed or separate molecules
administered which target at least two or an IGFBP, IGF-I and/or
IGF-I-receptor. Examples of suitable antisense molecules capable of
targetting the IGF-I receptor are those capable of interacting with
sequences selected from the list in Example 8. One particularly
useful antisense molecule is 5'-ATCTCTCCGCTTCCTTTC-3' (SEQ ID NO.
10). A particularly preferred embodiment of the present invention
contemplates a method of ameliorating the effects of psoriasis,
said method comprising contacting proliferating skin or skin
capable of proliferation with an effective amount of one or more
nucleic acid molecules or chemical analogues thereof capable of
inhibiting or otherwise reducing IGF-I mediated cell proliferation
wherein said one or more molecules comprises a polynucleotide
capable of interacting with mRNA directed from two or more of an
IGF-I gene, an IGF-I receptor gene or a gene encoding an IGFBP such
as IGFBP-2 and/or IGFBP-3.
In accordance with one aspect of the present invention the nucleic
acid molecule is topically applied in aqueous solution or in
conjunction with a cream, ointment, oil or other suitable carrier
and/or diluent. A single application may be sufficient depending on
the severity or exigencies of the condition although more commonly,
multiple applications are required ranging from hourly,
multi-hourly, daily, multi-daily, weekly or monthly, or in some
other suitable time interval. The treatment might comprise solely
the application of the nucleic acid molecule or this may be applied
in conjunction with other treatments for the skin proliferation
and/or inflammatory disorder being treated or for other associated
conditions including microbial infection, bleeding and the
formation of a variety of rashes.
As an alternative to or in conjunction with antisense therapy, the
subject invention extends to the nucleic acid molecule as, or
incorporating, a ribozyme including a minizyme to, for example,
IGF-I, its receptor or to molecules such as IGFBPs and in
particular IGFBP-2 and -3. Ribozymes are synthetic nucleic acid
molecules which possess highly specific endoribonuclease activity.
In particular, they comprise a hybridising region which is
complementary in nucleotide sequence to at least part of a target
RNA. Ribozymes are well described by Haseloff and Gerlach (8) and
in International Patent Application No. WO 89/05852. The present
invention extends to ribozymes which target mRNA specified by genes
encoding IGF-I, its receptor or one or more IGFBPs such as IGFBP-2
and/or IGFBP-3.
According to this embodiment, there is provided in a particularly
preferred aspect a ribozyme comprising a hybridising region and a
catalytic region wherein the hybridising region is capable of
hybridising to at least part of a target mRNA sequence transcribed
from a genomic gene corresponding to SEQ ID NO. 1 or SEQ ID NO. 2
wherein said catalytic domain is capable of cleaving said target
mRNA sequence to reduce or inhibit IGF-I mediated cell
proliferation and/or inflammation.
Yet another aspect of the present invention contemplates
co-suppression to reduce expression or to inhibit translation of an
endogenous gene encoding, for example, IGF-I, its receptor, or
IGFBPs such as IGFBP-2 and/or -3. In co-suppression, a second copy
of an endogenous gene or a substantially similar copy or analogue
of an endogenous gene is introduced into a cell following topical
administration. As with antisense molecules, nucleic acid molecules
defining a ribozyme or nucleic acid molecules useful in
co-suppression may first be protected such as by using a nonionic
backbone.
The efficacy of the nucleic acid molecules of the present invention
can be conveniently tested and screened using an in vitro system
comprising a basal keratinocyte cell line. A particularly useful
system comprises the HaCaT cell line described by Boukamp et al
(9). In one assay, IGF-I is added to an oligonucleotide treated
HaCaT cell line. Alternatively, growth of oligonucleotide treated
HaCaT cells is observed on a feeder layer of irradiated 3T3
fibroblasts. Using such in vitro assays, it is observed that
antisense oligonucleotides to IGFBP-3, for example, inhibit
production of IGFBP-3 by HaCaT cells. Other suitable animal models
include the nude mouse/human skin graft model (15; 16) and the
"flaky skin" mouse model (17; 18). In the nude mouse model,
microdermatome biopsies of psoriasis lesions are taken under local
anaesthetic from volunteers then transplanted to congenital athymic
(nude) mice. These transplanted human skin grafts maintain the
characteristic hyperproliferating epidermis for 6-8 weeks. They are
an established model for testing the efficacy of topically applied
therapies for psoriasis. In the "flaky skin" mouse model, the
fsn/fsn mutation produces mice with skin resembling human
psoriasis. This mouse, or another mutant mouse with a similar
phenotype is a further in vivo model to test the efficacy of
topically applied therapies for psoriasis.
Another aspect of the present invention contemplates a
pharmaceutical composition for topical administration which
comprises a nucleic acid molecule capable of inhibiting or
otherwise reducing IGF-I mediated cell proliferation such as
psoriasis and one or more pharmaceutically acceptable carriers
and/or diluents. Preferably, the nucleic acid molecule is an
antisense molecule to IGF-I, the IGF-I receptor or an IGFBP such as
IGFBP-2 and/or IGFBP-3 or comprises a ribozyme to one or more of
these targets or is a molecule suitable for co-suppression of one
or more of these targets. The composition may comprise a single
species of a nuleic acid molecule capable of targeting one of
IGF-I, its receptor or an IGFBP, such as IGFBP-2 or IGFBP-3 or may
be a multi-valent molecule capable of targeting two or more of
IGF-I, its receptor or an IGFBP, such as IGFBP-2 and/or
IGFBP-3.
The nucleic acid molecules may be administered in dispersions
prepared in creams, ointments, oil or other suitable carrier and/or
diluent such as glycerol, liquid polyethylene glycols and/or
mixtures thereof. Under ordinary conditions of storage and use,
these preparations may contain a preservative to prevent the growth
of microorganisms.
The pharmaceutical forms suitable for topical use include sterile
aqueous solutions (where water soluble) or dispersions and powders
for the extemporaneous preparation of topical solutions or
dispersion. In all cases, the form is preferably sterile although
this is not an absolute requirement and is stable under the
conditions of manufacture and storage. The carrier can be a solvent
or dispersion medium containing, for example, water, ethanol,
polyol (for example, glycerol, propylene glycol, and liquid
polyethylene glycol, and the like), suitable mixtures thereof and
vegetable oils. The proper fluidity can be maintained, for example,
by the use of a coating such as lecithin, by the maintenance of the
required particle size in the case of dispersion and by the use of
superfactants. The prevention of the action of microorganism can be
brought about by various antibacterial and antifungal agents, for
example, parabens, chlorobutanol, phenol, sorbic acid, thirmerosal
and the like. In many cases, it will be preferable to include
isotonic agents, for example, sugars or sodium chloride.
Topical solutions are prepared by incorporating the nucleic acid
molecule compound in the required amount in the appropriate solvent
with various of the other ingredients enumerated above, as
required, followed by where necessary filter sterilization.
As used herein "pharmaceutically acceptable carriers and/or
diluents" include any and all solvents, dispersion media, aqueous
solutions, coatings, antibacterial and antifungal agents, isotonic
and absorption delaying agents, and the like. The use of such media
and agents for pharmaceutical active substances is well known in
the art. Except insofar as any conventional media or agent is
incompatable with the active ingredient, use thereof in the
pharmaceutical compositions is contemplated. Supplementary active
ingredients can also be incorporated into the compositions.
Conveniently, the nucleic acid molecules of the present invention
are stored in freeze-dried form and are reconstituted prior to
use.
Yet another aspect of the present invention contemplates the use of
a nucleic acid molecule in the manufacture of a medicament for the
treatment of proliferative and/or inflammatory skin disorders
mediated by a growth factor. The proliferative and/or inflammatory
skin disorder is generally psoriasis and the nucleic acid molecule
targets IGF-I, the IGF-I receptor and/or an IGFBP such as IGFBP-2
and/or IGFBP-3.
Still a further aspect of the present invention contemplates an
agent comprising a nucleic acid molecule as hereinbefore defined
useful in the treatment of proliferative and/or inflammatory skin
disorders, such as psoriasis.
The present invention is further described by the following
non-limiting Figures and/or Examples.
DESCRIPTION OF THE DRAWINGS
FIG. 1 is a representation of the nucleotide sequence of
IGFBP-2(SEQ ID NO:1).
FIG. 2 is a representation of the nucleotide sequence of
IGFBP-3(SEQ ID NO:2).
FIG. 3 is a representation of the nucleotide sequence of
IGF-1-receptor (SEQ ID NO:3).
FIG. 4A is a photographic representation of a Western ligand blot
of HaCaT conditioned medium showing IGFBP-3 secreted in 24 hours
after 7 day treatment with phosphorothioate oligonucleotides
(BP3AS2, BP3AS3 and BP3S) at 0.5 .mu.M and 5 .mu.M; * no
oligonucleotide added.
FIG. 4B is a graphical representation of a scanning imaging
desitometry of Western ligand blot (FIG. 4A), showing relative band
intensities of IGFBP-3 and the 24kDa IGFBP-4 after treatment with
phosphorothioate oligonucleotides; * no oligonucleotide added.
FIG. 5A is a photographic representation of a Western ligand blot
of HaCaT conditioned medium showing IGFBP-3 secreted in 24 hours
after 7 day treatment with phosophorothioate oligonucleotide BP3AS2
at 0.5 .mu.M compared with several control oligonucleotides at 0.5
.mu.M. (a) oligonucleotide BP3AS2NS; (b) oligonucleotide BP3AS4;
(c) oligonucleotide BP3AS4NS; and (untreated), no oligonucleotide
added.
FIG 5B is a graphical representation of a scanning imaging
densitometry of Western ligand blot (FIG. 5A), showing relative
band intensities of IGFBP-3 after treatment with phosphorothioate
oligonucleotides as in FIG. 5A, showing IGFBP-3 band intensities
expressed as a percentage of the average band intensity from
conditioned medium of cells not treated with oligonucleotide.
FIG. 6 is a graphical representation showing inhibition of IGF-I
binding by antisense oligonucleotides to IGF-I receptor. IGFR.AS:
antisense; IGFR.S: sense.
FIG. 7 is a graphical representation showing inhibition of IGFBP-3
production in culture medium following initial treatment with
antisense oligonucleotides once daily over a 2 day period.
FIG. 8 is a graphical representation showing optimization of
IGFBP-3 antisense oligonucleotide concentration as determined by
relative IGFBP-3 concentration in culture medium.
EXAMPLE 1
IN VITRO ASSAY: CELLS
The differentiated human keratinocyte cell line, HaCaT (9) was used
in the in vitro assay. Cells at passage numbers 33 to 36 were
maintained as monolayer cultures in 5% v/v CO.sub.2 at 37.degree.
C. in Keratinocyte-SFM (Gibco) containing EGF and bovine pituitary
extract as supplied. Media containing foetal calf serum were
avoided because of the high content of IGF-I binding proteins in
serum.
Feeder layer plates of lethally irradiated 3T3 fibroblasts were
prepared exactly as described by Rheinwald and Green (10).
EXAMPLE 2
IN VITRO ASSAY: THYMIDINE INCORPORATION ASSAY
Cells were grown to 4 days post confluence in 2 cm.sup.2 wells with
daily medium changes of Keratinocyte-SFM, then the medium was
changed to DMEM (Cytosystems, Australia), with the following
additions: 25 mM Hepes, 0.19% w/v, sodium bicarbonate, 0.03% w/v
glutamine (Sigma Chemical Co, U.S.A.), 50 IU/ml penicillin and 50
.mu.g/ml streptomycin (Flow Laboratories). After 24 hours, IGF-I or
tIGF-I was added to triplicate wells, at the concentrations
indicated, in 0.5 ml fresh DMEM containing 0.02% v/v bovine serum
albumin (Sigma molecular biology grade) and incubated for a further
21 hours. [.sup.3 H]-Thymidine (0.1 .mu.Ci/well) was then added and
the cells incubated for a further 3 hours. The medium was then
aspirated and the cells washed once with ice-cold PBS and twice
with ice-cold 10% v/v TCA. The TCA-precipitated monolayers were
then solubilized with 0.25M NaOH (200 .mu.l/well), transferred to
scintillation vials and radioactivity determined by liquid
scintillation counting (Pharmacia Wallac 1410 liquid scintillation
counter).
EXAMPLE 3
WESTERN LIGAND BLOTTING
HaCaT conditioned medium (250 .mu.l) was concentrated by adding 750
.mu.l cold ethanol, incubating at -20.degree. C. for 2 hours and
centrifuging at 16,000 g for 20 min at 4.degree. C. The resulting
pellet was air dried, resuspended thoroughly in non-reducing
Laemmli sample buffer, heated to 90.degree. C. for 5 minutes and
separated on 12% w/v SDS-PAGE according to the method of Laemmli
(1970). Separated proteins were electrophoretically transferred to
nitrocellulose membrane (0.45 mm, Schleicher and Schuell, Dassel,
Germany) in a buffer containing 25 mM Tris, 192 mM glycine and 20%
v/v methanol. IGFBPs were then visualised by the procedure of
Hossenlopp et al (11), using [.sup.125 I]-IGF-I, followed by
autoradiography. Autoradiographs were scanned in a BioRad Model
GS-670 Imaging Densitometer and band densities were determined
using the Molecular Analyst program.
EXAMPLE 4
ANTISENSE OLIGONUCLEOTIDES
Phosphorothioate oligodeoxynucleotides were synthesised by
Bresatec, Adelaide, South Australia, Australia. The following
antisense sequences were used: BP3AS2, 5'-GCG CCC GCT GCA TGA CGC
CTG CAA C-3' (SEQ ID NO. 4), a 25 mer complementary to the start
codon region of the human IGFBP-3 mRNA; BP3AS3, 5'-CGG GCG GCT CAC
CTG GAG CTG GCG-3' (SEQ ID NO. 5), a 24 mer complementary to the
exon 1/intron 1 splice site; BP3AS4, 5'-AGG CGG CTG ACG GCA CTA-3'
(SEQ ID NO. 6), an 18 mer complementary to a region of the coding
sequence lacking RNA secondary structure and oligonucleotide-dimer
formation (using the computer software "OLIGO for PC"). Since
BP3AS4 was found to be ineffective at inhibiting IGFBP-3 synthesis,
it was used as a control. The following additional control
oligonucleotide sequences were used: BP3S, 5'-CAG GCG TCA TGC AGC
GGG C-3' (SEQ ID NO. 7), an 18 mer sense control sequence
equivalent to the start codon region; BP3AS2NS, 5'-CGG AGA TGC CGC
ATG CCA GCG CAG G-3' (SEQ ID NO. 8), a 25 mer randomised sequence
with the same GC content as BP3AS2; BP3AS4NS, 5'-GAC AGC GTC GGA
GCG ATC-3' (SEQ ID NO. 9), an 18 mer randomised sequence with the
same GC content as BP3AS4NS. Design of the oligonucleotides was
based on the human IGFBP-3 cDNA sequence of Spratt et al (12).
Cells were grown to one day post confluence in 2 cm.sup.2 wells
with daily medium changes of 0.5 ml Keratinocyte-SFM, then
subjected to daily medium changes of Keratinocyte-SFM for a further
4 days. Daily additions of 0.5 ml fresh Keratinocyte-SFM were then
continued for a further 7 days, except that at the time of medium
addition, 5 .mu.l oligonucleotide in PBS was added to give the
final concentrations indicated, then the wells were shaken to mix
the oligonucleotide. After the final addition, cells were incubated
for 24 hours and the medium collected for assay of IGFBPs. Cells
were then counted after trypsinisation in a Coulter Industrial D
Counter, Coulter Bedfordshire, UK. Cell numbers after
oligonucleotide treatment differed by less than 10%.
EXAMPLE 5
ANTISENSE OLIGONUCLEOTIDES INHIBIT IGFBP-3 SYNTHESIS
HaCaT cells secrete mainly IGFBP-3 (>95%), with the only other
IGFBP detectable in HaCaT conditioned medium being IGFBP-4
(<5%). The effect on IGFBP-3 and IGFBP-4 synthesis of antisense
oligonucleotides at two concentrations, 5 .mu.M and 0.5 .mu.M, was
tested. Two oligonucleotides were used, BP3AS2 and BP3AS3, directed
against the start site and the intron 1/exon 1 splice site,
respectively of the IGFBP-3 mRNA. As a control, a sense
oligonucleotide corresponding to the start site was used. As shown
in FIGS. 4A and 4B, all oligonucleotides at 5 .mu.M caused a
significant reduction of IGFBP-3 synthesis compared with untreated
cells, however, the two antisense oligonucleotides inhibited
IGFBP-3 synthesis of approximately 50% compared to the sense
control (FIG. 4B). The antisense oligonucleotide directed to the
start codon appeared to be more effective of the two, the
difference being more apparent at the lower concentration of 0.5
.mu.M. The cells of IGFBP-4 secreted by the HaCaT cells make
photographic reproduction of the bands on Western ligand blots
difficult, however densitometry measurements provide adequate
relative quantitation. This resulted in the significant observation
that IGFBP-4 levels were unaffected by oligonucleotide addition to
the cells, suggesting that the observed inhibitory effects on
IGFBP-3 are specific.
To further investigate the inhibitory effects of the more effective
of the two antisense oligonucleotides, BP3AS2, inhibition by this
oligonucleotide at 0.5 .mu.M was compared with a number of control
oligonucleotides, including one antisense oligonucleotide to
IGFBP-3 that had proved to be ineffective at 0.5 .mu.M. As shown in
FIGS. 5A and 5B, BP3AS2 was again inhibitory, resulting in levels
of IGFBP-3 of approximately 50% of the most non-specifically
inhibitory control oligonucleotide, the randomised equivalent of
BP3AS2. The other control oligonucleotides caused no reduction in
IGFBP-3 levels at 0.5 .mu.M, compared to untreated cells. Of
possible significance is the fact that this control
oligonucleotide, BP3AS2NS, like BP3AS2 itself, has the highest
potential T.sub.m of the three control oligonucleotides used in
this experiment, enhancing the probability of non-specific base
pairing with non-target mRNAs. However, the lack of inhibition of
IGFBP-4 secretion by BP3AS2 suggests that this oligonucleotide is
selective even compared with the most closely related protein
likely to be present in this cell line.
EXAMPLE 6
ANTISENSE OLIGONUCLEOTIDES OF IGFBP2
Antisense oligonucleotides to IGFBP2 may be selected from molecules
capable of interacting with one or more oligonucleotides selected
from oligonucleotides having the sequence of nucleotides 1-15,
2-16, 3-17, 4-18, 5-19, 6-20, 7-21, 8-22, 9-23, 10-24, 11-25,
12-26, 13-27, 14-28, 15-29, 16-30, 17-31, 18-32, 19-33, 20-34,
21-35, 22-36, 23-37, 24-38, 25-39, 26-40, 27-41, 28-42, 29-43,
30-44, 31-45, 32-46, 33-47, 34-48, 35-49, 36-50, 37-51, 38-52,
39-53, 40-54, 41-55, 42-56, 43-57, 44-58, 45-59, 46-60, 47-61,
48-62, 49-63, 50-64, 51-65, 52-66, 53-67, 54-68, 55-69, 56-70,
57-71, 58-72, 59-73, 60-74, 61-75, 62-76, 63-77, 64-78, 65-79,
66-80, 67-81, 68-82, 69-83, 70-84, 71-85, 72-86, 73-87, 74-88,
75-89, 76-90, 77-91, 78-92, 79-93, 80-94, 81-95, 82-96, 83-97,
84-98, 85-99, 86-100, 87-101, 88-102, 89-103, 90-104, 91-105,
92-106, 93-107, 94-108, 95-109, 96-110, 97-111, 98-112, 99-113,
100-114, 101-115, 102-116, 103-117, 104-118, 105-119, 106-120,
107-121, 108-122, 109-123, 110-124, 111-125, 112-126, 113-127,
114-128, 115-129, 116-130, 117-131, 118-132, 119-133, 120-134,
121-135, 122-136, 123-137, 124-138, 125-139, 126-140, 127-141,
128-142, 129-143, 130-144, 131-145, 132-146, 133-147, 134-148,
135-149, 136-150, 137-151, 138-152, 139-153, 140-154, 141-155,
142-156, 143-157, 144-158, 145-159, 146-160, 147-161 148-162,
149-163, 150-164, 151-165, 152-166, 153-167, 154-168, 155-169,
156-170, 157-171, 158-172, 159-173, 160-174, 161-175,162-176,
163-177, 164-178, 165-179, 166-180, 167-181, 168-182, 169-183,
170-184, 171-185, 172-186, 173-187, 174-188, 175-189, 176-190,
177-191, 178-192, 179-193, 180-194, 181-195, 182-196, 183-197,
184-198, 185-199, 186-200, 187-201, 188-202, 189-203, 190-204,
191-205, 192-206, 193-207, 194-208, 195-209, 196-210, 197-211,
198-212, 199-213, 200-214, 201-215, 202-216, 203-217, 204-218,
205-219, 206-220, 207-221, 208-222, 209-223, 210-224, 211-225,
212-226, 213-227, 214-228, 215-229, 216-230, 217-231, 218-232,
219-233, 220-234, 221-235, 222-236, 223-237, 224-238, 225-239,
226-240, 227-241, 228-242, 229-243, 230-244, 231-245, 232-246,
233-247, 234-248, 235-249, 236-250, 237-251, 238-252, 239-253,
240-254, 241-255, 242-256, 243-257, 244-258, 245-259, 246-260,
247-261, 248-262, 249-263, 250-264, 251-265, 252-266, 253-267,
254-268, 255-269, 256-270, 257-271, 258-272, 259-273, 260-274,
261-275, 262-276, 263-277, 264-278, 265-279, 266-280, 267-281,
268-282, 269-283, 270-284, 271-285, 272-286, 273-287, 274-288,
275-289, 276-290, 277-291, 278-292, 279-293, 280-294, 281-295,
282-296, 283-297, 284-298, 285-299, 286-300, 287-301, 288-302,
289-303, 290-304, 291-305, 292-306, 293-307, 294-308, 295-309,
296-310, 297-311, 298-312, 299-313, 300-314, 301-315, 302-315,
303-317, 304-318, 305-319, 306-320, 307-321, 308-322, 309-323,
310-324, 311-325, 312-326, 313-327, 314-328, 315-329, 316-330,
317-331, 318-332, 319-333, 320-334, 321-335, 322-336, 323-337,
324-338, 325-339, 326-340, 327-341, 328-342, 329-343, 330-344,
331-345, 332-346, 333-347, 334-348, 335-349, 336-350, 337-351,
338-352, 339-353, 340-354, 341-355, 342-356, 343-357, 344-358,
345-359, 346-360, 347-361, 348-362, 349-363, 350-364, 351-365,
352-366, 353-367, 354-368, 355-369, 356-370, 357-371, 358-372,
359-373, 360-374, 361-375, 362-376, 363-377, 364-378, 365-379,
366-380, 367-381, 368-382, 369-383, 370-384, 371-385, 372-386,
373-387, 374-388, 375-389, 376-390, 377-391, 378-392, 379-393,
380-394, 381-395, 382-396, 383-397, 384-398, 385-399, 386-400,
387-401, 388-402, 389-403, 390-404, 391-405, 392-406, 393-407,
394-408, 395-409, 396-410, 397-411, 398-412, 399-413, 400-414,
401-415, 402-416, 403-417, 404-418, 405-419, 406-420, 407-421,
408-422, 409-423, 410-424, 411-425, 412-426, 413-427, 414-428,
415-429, 416-430, 417-431, 418-432, 419-433, 420-434, 421-435,
422-436, 423-437, 424-438, 425-439, 426-440, 427-441, 428-442,
429-443, 430-444, 431-445, 432-446, 433-447, 434-448, 435-449,
436-450, 437-451, 438-452, 439-453, 440-454, 441-455, 442-456,
443-457, 444-458, 445-459, 446-460, 447-461, 448-462, 449-463,
450-464, 451-465, 452-466, 453-467, 454-468, 455-469, 456-470,
457-471, 458-472, 459-473, 460-474, 461-475, 462-476, 463-477,
464-478, 465-479, 466-480, 467-481, 468-482, 469-483, 470-484,
471-485, 472-486, 473-487, 474-488, 475-489, 476-490, 477-491,
478-492, 479-493, 480-494, 481-495, 482-496, 483-497, 484-498,
485-499, 486-500, 487-501, 488-502, 489-503, 490-504, 491-505,
492-506, 493-507, 494-508, 495-509, 496-510, 497-511, 498-513,
499-514, 500-515, 501-515, 502-516, 503-517, 504-518, 505-519,
506-520, 507-521, 508-522, 509-523, 510-524, 511-525, 512-526,
513-527, 514-528, 515-529, 516-530, 517-531, 518-532, 519-533,
520-534, 521-535, 522-536, 523-537, 524-538, 525-539, 526-540,
527-541, 528-542, 529-543, 530-544, 531-545, 532-546, 533-547,
534-548, 535-549, 536-550, 537-551, 538-552, 539-553, 540-554,
541-555, 542-556, 543-557, 544-558, 545-559, 546-560, 547-561,
548-562, 549-563, 550-564, 551-565, 552-566, 553-567, 554-568,
555-569, 556-570, 557-571, 558-572, 559-573, 560-574, 561-575,
562-576, 563-577, 564-578, 565-579, 566-580, 567-581, 568-582,
569-583, 570-584, 571-585, 572-586, 573-587, 574-588, 575-589,
576-590, 577-591, 578-592, 579-593, 580-594, 581-595, 582-596,
583-597, 584-598, 585-599, 586-600, 587-601, 588-602, 589-603,
590-604, 591-605, 592-606, 593-607, 594-608, 595-609, 596-610,
597-611, 598-612, 599-613, 600-614, 601-615, 602-616, 603-617,
604-618, 605-619, 606-620, 607-621, 608-622, 609-623, 610-624,
611-625, 612-626, 613-627, 614-628, 615-629, 616-630, 617-631,
618-632, 619-633, 620-634, 621-635, 622-636, 623-637, 624-638,
625-639, 626-640, 627-641, 628-642, 629-643, 630-644, 631-645,
632-646, 633-647, 634-648, 635-649, 636-650, 637-651, 638-652,
639-653, 640-654, 641-655, 642-656, 643-657, 644-658, 645-659,
646-660, 647-661, 648-662, 649-663, 650-664, 651-665, 652-666,
653-667, 654-668, 655-669, 656-670, 657-671, 658-672, 659-673,
660-674, 661-675, 662-676, 663-677, 664-678, 665-679, 666-680,
667-681, 668-682, 669-683, 670-684, 671-685, 672-686, 673-687,
674-688, 675-689, 676-690, 677-691, 678-692, 679-693, 680-694,
681-695, 682-696, 683-697, 684-698, 685-699, 686-700, 687-701,
688-702, 689-703, 690-704, 691-705, 692-706, 693-707, 694-708,
695-709, 696-710, 697-711, 698-712, 699-713, 700-714, 701-715,
702-716, 703-717, 704-718, 705-719, 706-720, 707-721, 708-722,
709-723, 710-724, 711-725, 712-726, 713-727, 714-728, 715-729,
716-730, 717-731, 718-732, 719-733, 720-734, 721-735, 722-736,
723-737, 724-738, 725-739, 726-740, 727-741, 728-742, 729-743,
730-744, 731-745, 732-746, 733-747, 734-748, 735-749, 736-750,
737-751, 738-752, 739-753, 740-754, 741-755, 742-756, 743-757,
744-758, 745-759, 746-760, 747-761, 748-762, 749-763, 750-764,
751-765, 752-766, 753-767, 754-768, 755-769, 756-770, 757-771,
758-772, 759-773, 760-774, 761-775, 762-776, 763-777, 764-778,
765-779, 766-780, 767-781, 768-782, 769-783, 770-784, 771-785,
772-786, 773-787, 774-788, 775-789, 776-790, 777-791 778-792,
779-793, 780-794, 781-795, 782-796, 783-797, 784-798, 785-799,
786-800, 787-801, 788-802, 789-803, 790-804, 791-805, 792-806,
793-807, 794-808, 795-809, 796-810, 797-811, 798-812, 799-813,
800-814, 801-815, 802-816, 803-817, 804-818, 805-819, 806-820,
807-821, 808-822, 809-823, 810-824, 811-825, 812-826, 813-827,
814-828, 815-829, 816-830, 817-831, 818-832, 819-833, 820-834,
821-835, 822-836, 823-837, 824-838, 825-839, 826-840, 827-841,
828-842, 829-843, 830-844, 831-845, 832-846, 833-847, 834-848,
835-849, 836-850, 837-851, 838-852, 839-853, 840-854, 841-855,
842-856, 843-857, 844-858, 845-859, 846-860, 847-861, 848-862,
849-863, 850-864, 851-865, 852-866, 853-867, 854-868, 855-869,
856-870, 857-871, 858-872, 859-873, 860-874, 861-875, 862-876,
863-877, 864-878, 865-879, 866-880, 867-881, 868-882, 869-883,
870-884, 871-885, 872-886, 873-887, 874-888, 875-889, 876-890,
877-891, 878-892, 879-893, 880-894, 881-895, 882-896, 883-897,
884-898, 885-899, 886-900, 887-901, 888-902, 889-903, 890-904,
891-905, 892-906, 893-907, 894-908, 895-909, 896-910, 897-911,
898-912, 899-913, 900-914, 901-915, 902-916, 903-917, 904-918,
905-919, 906-920, 907-921, 908-922, 909-923, 910-924, 911-925,
912-926, 913-927, 914-928, 915-929, 916-930, 917-931, 918-932,
919-933, 920-934, 921-935, 922-936, 923-937, 924-938, 925-939,
926-940, 927-941, 928-942, 929-943, 930-944, 931-945, 932-946,
933-947, 934-948, 935-949, 936-950, 937-951, 938-952, 939-953,
940-954, 941-955, 942-956, 943-957, 944-958, 945-959, 946-960,
947-961, 948-962, 949-963, 950-964, 951-965, 952-966, 953-967,
954-968, 955-969, 956-970, 957-971, 958-972, 959-973, 960-974,
961-975, 962-976, 963-977, 964-978, 965-979, 966-980, 967-981,
968-982, 969-983, 970-984, 971-985, 972-986, 973-987, 974-988,
975-989, 976-990, 977-991, 978-992, 979-993, 980-994, 981-995,
982-996, 983-997, 984-998, 985-999, 986-1000, 987-1001, 988-1002,
989-1003, 990-1004, 991-1005, 992-1006, 993-1007, 994-1008,
995-1009, 996-1010, 997-1011, 998-1012, 999-1013, 1000-1014,
1001-1015, 1002-1016, 1003-1017, 1004-1018, 1005-1019, 1006-1020,
1007-1021, 1008-1022, 1009-1023, 1010-1024, 1011-1025, 1012-1026,
1013-1027, 1014-1028, 1015-1029, 1016-1030, 1017-1031, 1018-1032,
1019-1033, 1020-1034, 1021-1035, 1022-1036, 1023-1037, 1024-1038,
1025-1039, 1026-1040, 1027-1041, 1028-1042, 1029-1043, 1030-1044,
1031-1045, 1032-1046, 1033-1047, 1034-1048, 1035-1049, 1036-1050,
1037-1051, 1038-1052, 1039-1053, 1040-1054, 1041-1055, 1042-1056,
1043-1057, 1044-1058, 1045-1059, 1046-1060, 1047-1061, 1048-1062,
1049-1063, 1050-1064, 1051-1065, 1052-1066, 1053-1067, 1054-1068,
1055-1069, 1056-1070, 1057-1071, 1058-1072, 1059-1073, 1060-1074,
1061-1075, 1062-1076, 1063-1077, 1064-1078, 1065-1079, 1066-1080,
1067-1081, 1068-1082, 1069-1083, 1070-1084, 1071-1085, 1072-1086
1073-1087, 1074-1088, 1075-1089, 1076-1090, 1077-1091, 1078-1092,
1079-1093, 1080-1094, 1081-1095, 1082-1096, 1083-1097, 1084-1098,
1085-1099, 1086-1100, 1087-1101, 1088-1102, 1089-1103, 1090-1104,
1091-1105, 1092-1106, 1093-1107, 1094-1108, 1095-1109, 1096-1110,
1097-1111, 1098-1112, 1099-1113, 1100-1114, 1101-1115, 1102-1116,
1103-1117, 1104-1118, 1105-1119, 1106-1120, 1107-1121, 1108-1122,
1109-1123, 1110-1124, 1111-1125, 1112-1126, 1113-1127, 1114-1128,
1115-1129, 1116-1130, 1117-1131, 1118-1132, 1119-1133, 1120-1134,
1121-1135, 1122-1136, 1123-1137, 1124-1138, 1125-1139, 1126-1140,
1127-1141, 1128-1142, 1129-1143, 1130-1144, 1131-1145, 1132-1146,
1133-1147, 1134-1148, 1135-1149, 1136-1150, 1137-1151, 1138-1152,
1139-1153, 1140-1154, 1141-1155, 1142-1156, 1143-1157, 1144-1158,
1145-1159, 1146-1160, 1147-1161, 1148-1162, 1149-1163, 1150-1164,
1151-1165, 1152-1166, 1153-1167, 1154-1168, 1155-1169, 1156-1170,
1157-1171, 1158-1172, 1159-1173, 1160-1174, 1161-1175, 1162-1176,
1163-1177, 1164-1178, 1165-1179, 1166-1180, 1167-1181, 1168-1182,
1169-1183, 1170-1184, 1171-1185, 1172-1186, 1173-1187, 1174-1188,
1175-1189, 1176-1190, 1177-1191, 1178-1192, 1179-1193, 1180-1194,
1181-1195, 1182-1196, 1183-1197, 1184-1198, 1185-1199, 1186-1200,
1187-1201, 1188-1202, 1189-1203, 1190-1204, 1191-1205, 1192-1206,
1193-1207, 1194-1208, 1195-1209, 1196-1210, 1197-1211, 1198-1212,
1199-1213, 1200-1214, 1201-1215, 1202-1216, 1203-1217, 1204-1218,
1205-1219, 1206-1220, 1207-1221, 1208-1222, 1209-1223, 1210-1224,
1211-1225, 1212-1226, 1213-1227, 1214-1228, 1215-1229, 1216-1230,
1217-1231, 1218-1232, 1219-1233, 1220-1234, 1221-1235, 1222-1236,
1223-1237, 1224-1238, 1225-1239, 1226-1240, 1227-1241, 1228-1242,
1229-1243, 1230-1244, 1231-1245, 1232-1246, 1233-1247, 1234-1248,
1235-1249, 1236-1250, 1237-1251, 1238-1252, 1239-1253, 1240-1254,
1241-1255, 1242-1256, 1243-1257, 1244-1258, 1245-1259, 1246-1260,
1247-1261, 1248-1262, 1249-1263, 1250-1264, 1251-1265, 1252-1266,
1253-1267, 1254-1268, 1255-1269, 1256-1270, 1257-1271, 1258-1272,
1259-1273, 1260-1274, 1261-1275, 1262-1276, 1263-1277, 1264-1278,
1265-1279, 1266-1280, 1267-1281, 1268-1282, 1269-1283, 1270-1284,
1271-1285, 1272-1286, 1273-1287, 1274-1288, 1275-1289, 1276-1290,
1277-1291, 1278-1292, 1279-1293, 1280-1294, 1281-1295, 1282-1296,
1283-1297, 1284-1298, 1285-1299, 1286-1300, 1287-1301, 1288-1302,
1289-1303, 1290-1304, 1291-1305, 1292-1306, 1293-1307, 1294-1308,
1295-1309, 1296-1310, 1297-1311, 1298-1312, 1299-1313, 1300-1314,
1301-1315, 1302-1315, 1303-1317, 1304-1318, 1305-1319, 1306-1320,
1307-1321, 1308-1322, 1309-1323, 1310-1324, 1311-1325, 1312-1326,
1313-1327, 1314-1328, 1315-1329, 1316-1330, 1317-1331, 1318-1332,
1319-1333, 1320-1334, 1321-1335, 1322-1336, 1323-1337, 1324-1338,
1325-1339, 1326-1340, 1327-1341, 1328-1342, 1329-1343, 1330-1344,
1331-1345, 1332-1346, 1333-1347, 1334-1348, 1335-1349, 1336-1350,
1337-1351, 1338-1352, 1339-1353, 1340-1354, 1341-1355, 1342-1356,
1343-1357, 1344-1358, 1345-1359, 1346-1360, 1347-1361, 1348-1362,
1349-1363, 1350-1364, 1351-1365, 1352-1366, 1353-1367, 1354-1368,
1355-1369, 1356-1370, 1357-1371, 1358-1372, 1359-1373, 1360-1374,
1361-1375, 1362-1376, 1363-1377, 1364-1378, 1365-1379, 1366-1380,
1367-1381, 1368-1382, 1369-1383, 1370-1384, 1371-1385, 1372-1386,
1373-1387, 1374-1388, 1375-1389, 1376-1390, 1377-1391, 1378-1392,
1379-1393, 1380-1394, 1381-1395, 1382-1396, 1383-1397, 1384-1398,
1385-1399, 1386-1400, 1387-1401, 1388-1402, 1389-1403, 1390-1404,
1391-1405, 1392-1406, 1393-1407, 1394-1408, 1395-1409, 1396-1410,
1397-1411, 1398-1412, 1399-1413, 1400-1414, 1401-1415, 1402-1416,
1403-1417, 1404-1418, 1405-1419, 1406-1420, 1407-1421, 1408-1422,
1409-1423, 1410-1424, 1411-1425, 1412-1426, 1413-1427, 1414-1428,
1415-1429, 1416-1430, 1417-1431, 1418-1432, and 1419-1433 of SEQ ID
NO:1.
EXAMPLE 7
ANTISENSE OLIGONUCLEOTIDES OF IGFBP3
Antisense oligonucleotides to IGFBP3 may be selected from molecules
capable of interacting with one or more oligonucleotides selected
from oligonucleotides having the sequence of nucleotides 1-15,
2-16, 3-17, 4-18, 5-19, 6-20, 7-21, 8-22, 9-23, 10-24, 11-25,
12-26, 13-27, 14-28, 15-29, 16-30, 17-31, 18-32, 19-33, 20-34,
21-35, 22-36, 23-37, 24-38, 25-39, 26-40, 27-41, 28-42, 29-43,
30-44, 31-45, 32-46, 33-47, 34-48, 35-49, 36-50, 37-51, 38-52,
39-53, 40-54, 41-55, 42-56, 43-57, 44-58, 45-59, 46-60, 47-61,
48-62, 49-63, 50-64, 51-65, 52-66, 53-67, 54-68, 55-69, 56-70,
57-71, 58-72, 59-73, 60-74, 61-75, 62-76, 63-77, 64-78, 65-79,
66-80, 67-81, 68-82, 69-83, 70-84, 71-85, 72-86, 73-87, 74-88,
75-89, 76-90, 77-91, 78-92, 79-93, 80-94, 81-95, 82-96, 83-97,
84-98, 85-99, 86-100, 87-101, 88-102, 89-103, 90-104, 91-105,
92-106, 93-107, 94-108, 95-109, 96-110, 97-111, 98-112, 99-113,
100-114, 101-115, 102-116, 103-117, 104-118, 105-119, 106-120,
107-121, 108-122, 109-123, 110-124, 111-125, 112-126, 113-127,
114-128, 115-129, 116-130, 117-131, 118-132, 119-133, 120-134,
121-135, 122-136, 123-137, 124-138, 125-139, 126-140, 127-141,
128-142, 129-143, 130-144, 131-145, 132-146, 133-147, 134-148,
135-149, 136-150, 137-151, 138-152, 139-153, 140-154, 141-155,
142-156, 143-157, 144-158, 145-159, 146-160, 147-161, 148-162,
149-163, 150-164, 151-165, 152-166, 153-167, 154-168, 155-169,
156-170, 157-171, 158-172, 159-173, 160-174, 161-175, 162-176,
163-177, 164-178, 165-179, 166-180, 167-181, 168-182, 169-183,
170-184, 171-185, 172-186, 173-187, 174-188, 175-189, 176-190,
177-191, 178-192, 179-193, 180-194, 181-195, 182-196, 183-197,
184-198, 185-199, 186-200, 187-201, 188-202, 189-203, 190-204,
191-205, 192-206, 193-207, 194-208, 195-209, 196-210, 197-211,
198-212, 199-213, 200-214, 201-215, 202-216, 203-217, 204-218,
205-219, 206-220, 207-221, 208-222, 209-223, 210-224, 211-225,
212-226, 213-227, 214-228, 215-229, 216-230, 217-231, 218-232,
219-233, 220-234, 221-235, 222-236, 223-237, 224-238, 225-239,
226-240, 227-241, 228-242, 229-243, 230-244, 231-245, 232-246,
233-247, 234-248, 235-249, 236-250, 237-251, 238-252, 239-253,
240-254, 241-255, 242-256, 243-257, 244-258, 245-259, 246-260,
247-261, 248-262, 249-263, 250-264, 251-265, 252-266, 253-267,
254-268, 255-269, 256-270, 257-271, 258-272, 259-273, 260-274,
261-275, 262-276, 263-277, 264-278, 265-279, 266-280, 267-281,
268-282, 269-283, 270-284, 271-285, 272-286, 273-287, 274-288,
275-289, 276-290, 277-291, 278-292, 279-293, 280-294, 281-295,
282-296, 283-297, 284-298, 285-299, 286-300, 287-301, 288-302,
289-303, 290-304, 291-305, 292-306, 293-307, 294-308, 295-309,
296-310, 297-311, 298-312, 299-313, 300-314, 301-315, 302-315,
303-317, 304-318, 305-319, 306-320, 307-321, 308-322, 309-323,
310-324, 311-325, 312-326, 313-327, 314-328, 315-329, 316-330,
317-331, 318-332, 319-333, 320-334, 321-335, 322-336, 323-337,
324-338, 325-339, 326-340, 327-341, 328-342, 329-343, 330-344,
331-345, 332-346, 333-347, 334-348, 335-349, 336-350, 337-351,
338-352, 339-353, 340-354, 341-355, 342-356, 343-357, 344-358,
345-359, 346-360, 347-361, 348-362, 349-363, 350-364, 351-365,
352-366, 353-367, 354-368, 355-369, 356-370, 357-371, 358-372,
359-373, 360-374, 361-375, 362-376, 363-377, 364-378, 365-379,
366-380, 367-381, 368-382, 369-383, 370-384, 371-385, 372-386,
373-387, 374-388, 375-389, 376-390, 377-391, 378-392, 379-393,
380-394, 381-395, 382-396, 383-397, 384-398, 385-399, 386-400,
387-401, 388-402, 389-403, 390-404, 391-405, 392-406, 393-407,
394-408, 395-409, 396-410, 397-411, 398-412, 399-413, 400-414,
401-415, 402-416, 403-417, 404-418, 405-419, 406-420, 407-421,
408-422, 409-423, 410-424, 411-425, 412-426, 413-427, 414-428,
415-429, 416-430, 417-431, 418-432, 419-433, 420-434, 421-435,
422-436, 423-437, 424-438, 425-439, 426-440, 427-441, 428-442,
429-443, 430-444, 431-445, 432-446, 433-447, 434-448, 435-449,
436-450, 437-451, 438-452, 439-453, 440-454, 441-455, 442-456,
443-457, 444-458, 445-459, 446-460, 447-461, 448-462, 449-463,
450-464, 451-465, 452-466, 453-467, 454-468, 455-469, 456-470,
457-471, 458-472, 459-473, 460-474, 461-475, 462-476, 463-477,
464-478, 465-479, 466-480, 467-481, 468-482, 469-483, 470-484,
471-485, 472-486, 473-487, 474-488, 475-489, 476-490, 477-491,
478-492, 479-493, 480-494, 481-495, 482-496, 483-497, 484-498,
485-499, 486-500, 487-501, 488-502, 489-503, 490-504, 491-505,
492-506, 493-507, 494-508, 495-509, 496-510, 497-511, 498-513,
499-514, 500-515, 501-515, 502-516, 503-517, 504-518, 505-519,
506-520, 507-521, 508-522, 509-523, 510-524, 511-525, 512-526,
513-527, 514-528, 515-529, 516-530, 517-531, 518-532, 519-533,
520-534, 521-535, 522-536, 523-537, 524-538, 525-539, 526-540,
527-541, 528-542, 529-543, 530-544, 531-545, 532-546, 533-547,
534-548, 535-549, 536-550, 537-551, 538-552, 539-553, 540-554,
541-555, 542-556, 543-557, 544-558, 545-559, 546-560, 547-561,
548-562, 549-563, 550-564, 551-565, 552-566, 553-567, 554-568,
555-569, 556-570, 557-571, 558-572, 559-573, 560-574, 561-575,
562-576, 563-577, 564-578, 565-579, 566-580, 567-581, 568-582,
569-583, 570-584, 571-585, 572-586, 573-587, 574-588, 575-589,
576-590, 577-591, 578-592, 579-593, 580-594, 581-595, 582-596,
583-597, 584-598, 585-599, 586-600, 587-601, 588-602, 589-603,
590-604, 591-605, 592-606, 593-607, 594-608, 595-609, 596-610,
597-611, 598-612, 599-613, 600-614, 601-615, 602-616, 603-617,
604-618, 605-619, 606-620, 607-621, 608-622, 609-623, 610-624,
611-625, 612-626, 613-627, 614-628, 615-629, 616-630, 617-631,
618-632, 619-633, 620-634, 621-635, 622-636, 623-637, 624-638,
625-639, 626-640, 627-641, 628-642, 629-643, 630-644, 631-645,
632-646, 633-647, 634-648, 635-649, 636-650, 637-651, 638-652,
639-653, 640-654, 641-655, 642-656, 643-657, 644-658, 645-659,
646-660, 647-661, 648-662, 649-663, 650-664, 651-665, 652-666,
653-667, 654-668, 655-669, 656-670, 657-671, 658-672, 659-673,
660-674, 661-675, 662-676, 663-677, 664-678, 665-679, 666-680,
667-681, 668-682, 669-683, 670-684, 671-685, 672-686, 673-687,
674-688, 675-689, 676-690, 677-691, 678-692, 679-693, 680-694,
681-695, 682-696, 683-697, 684-698, 685-699, 686-700, 687-701,
688-702, 689-703, 690-704, 691-705, 692-706, 693-707, 694-708,
695-709, 696-710, 697-711, 698-712, 699-713, 700-714, 701-715,
702-716, 703-717, 704-718, 705-719, 706-720, 707-721, 708-722,
709-723, 710-724, 711-725, 712-726, 713-727, 714-728,715-729,
716-730, 717-731, 718-732, 719-733, 720-734, 721-735, 722-736,
723-737, 724-738, 725-739, 726-740, 727-741, 728-742, 729-743,
730-744, 731-745, 732-746, 733-747, 734-748, 735-749, 736-750,
737-751, 738-752, 739-753, 740-754, 741-755, 742-756, 743-757,
744-758, 745-759, 746-760, 747-761, 748-762, 749-763, 750-764,
751-765, 752-766, 753-767, 754-768, 755-769, 756-770, 757-771,
758-772, 759-773, 760-774, 761-775, 762-776, 763-777, 764-778,
765-779, 766-780, 767-781, 768-782, 769-783, 770-784, 771-785,
772-786, 773-787, 774-788, 775-789, 776-790, 777-791, 778-792,
779-793, 780-794, 781-795, 782-796, 783-797, 784-798, 785-799,
786-800, 787-801, 788-802, 789-803, 790-804, 791-805, 792-806,
793-807, 794-808, 795-809, 796-810, 797-811, 798-812, 799-813,
800-814, 801-815, 802-816, 803-817, 804-818, 805-819, 806-820,
807-821, 808-822, 809-823, 810-824, 811-825, 812-826, 813-827,
814-828, 815-829, 816-830, 817-831, 818-832, 819-833, 820-834,
821-835, 822-836, 823-837, 824-838, 825-839, 826-840, 827-841,
828-842, 829-843, 830-844, 831-845, 832-846, 833-847, 834-848,
835-849, 836-850, 837-851, 838-852, 839-853, 840-854, 841-855,
842-856, 843-857, 844-858, 845-859, 846-860, 847-861, 848-862,
849-863, 850-864, 851-865, 852-866, 853-867, 854-868, 855-869,
856-870, 857-871, 858-872, 859-873, 860-874, 861-875, 862-876,
863-877, 864-878, 865-879, 866-880, 867-881, 868-882, 869-883,
870-884, 871-885, 872-886, 873-887, 874-888, 875-889, 876-890,
877-891, 878-892, 879-893, 880-894, 881-895, 882-896, 883-897,
884-898, 885-899, 886-900, 887-901, 888-902, 889-903, 890-904,
891-905, 892-906, 893-907, 894-908, 895-909, 896-910, 897-911,
898-912, 899-913, 900-914, 901-915, 902-916, 903-917, 904-918,
905-919, 906-920, 907-921, 908-922, 909-923, 910-924, 911-925,
912-926, 913-927, 914-928, 915-929, 916-930, 917-931, 918-932,
919-933, 920-934, 921-935, 922-936, 923-937, 924-938, 925-939,
926-940, 927-941, 928-942, 929-943, 930-944, 931-945, 932-946,
933-947, 934-948, 935-949, 936-950, 937-951, 938-952, 939-953,
940-954, 941-955, 942-956, 943-957, 944-958, 945-959, 946-960,
947-961, 948-962, 949-963, 950-964, 951-965, 952-966, 953-967,
954-968, 955-969, 956-970, 957-971, 958-972, 959-973, 960-974,
961-975, 962-976, 963-977, 964-978, 965-979, 966-980, 967-981,
968-982, 969-983, 970-984, 971-985, 972-986, 973-987, 974-988,
975-989, 976-990, 977-991, 978-992, 979-993, 980-994, 981-995,
982-996, 983-997, 984-998, 985-999, 986-1000, 987-1001, 988-1002,
989-1003, 990-1004, 991-1005, 992-1006, 993-1007, 994-1008,
995-1009, 996-1010, 997-1011, 998-1012, 999-1013, 1000-1014,
1001-1015, 1002-1016, 1003-1017, 1004-1018, 1005-1019, 1006-1020,
1007-1021, 1008-1022, 1009-1023, 1010-1024, 1011-1025, 1012-1026,
1013-1027, 1014-1028, 1015-1029, 1016-1030, 1017-1031, 1018-1032,
1019-1033, 1020-1034, 1021-1035, 1022-1036, 1023-1037, 1024-1038,
1025-1039, 1026-1040, 1027-1041, 1028-1042, 1029-1043, 1030-1044,
1031-1045, 1032-1046, 1033-1047, 1034-1048, 1035-1049, 1036-1050,
1037-1051, 1038-1052, 1039-1053, 1040-1054, 1041-1055, 1042-1056,
1043-1057, 1044-1058, 1045-1059, 1046-1060, 1047-1061, 1048-1062,
1049-1063, 1050-1064, 1051-1065, 1052-1066, 1053-1067, 1054-1068,
1055-1069, 1056-1070, 1057-1071, 1058-1072, 1059-1073, 1060-1074,
1061-1075, 1062-1076, 1063-1077, 1064-1078, 1065-1079, 1066-1080,
1067-1081, 1068-1082, 1069-1083, 1070-1084, 1071-1085, 1072-1086,
1073-1087, 1074-1088, 1075-1089, 1076-1090, 1077-1091, 1078-1092,
1079-1093, 1080-1094, 1081-1095, 1082-1096, 1083-1097, 1084-1098,
1085-1099, 1086-1100, 1087-1101, 1088-1102, 1089-1103, 1090-1104,
1091-1105, 1092-1106, 1093-1107, 1094-1108, 1095-1109, 1096-1110,
1097-1111, 1098-1112, 1099-1113, 1100-1114, 1101-1115, 1102-1116,
1103-1117, 1104-1118, 1105-1119, 1106-1120, 1107-1121, 1108-1122,
1109-1123, 1110-1124, 1111-1125, 1112-1126, 1113-1127, 1114-1128,
1115-1129, 1116-1130, 1117-1131, 1118-1132, 1119-1133, 1120-1134,
1121-1135, 1122-1136, 1123-1137, 1124-1138, 1125-1139, 1126-1140,
1127-1141, 1128-1142, 1129-1143, 1130-1144, 1131-1145, 1132-1146,
1133-1147, 1134-1148, 1135-1149, 1136-1150, 1137-1151, 1138-1152,
1139-1153, 1140-1154, 1141-1155, 1142-1156, 1143-1157, 1144-1158,
1145-1159, 1146-1160, 1147-1161, 1148-1162, 1149-1163, 1150-1164,
1151-1165, 1152-1166, 1153-1167, 1154-1168, 1155-1169, 1156-1170,
1157-1171, 1158-1172, 1159-1173, 1160-1174, 1161-1175, 1162-1176,
1163-1177, 1164-1178, 1165-1179, 1166-1180, 1167-1181, 1168-1182,
1169-1183, 1170-1184, 1171-1185, 1172-1186, 1173-1187, 1174-1188,
1175-1189, 1176-1190, 1177-1191, 1178-1192, 1179-1193, 1180-1194,
1181-1195, 1182-1196, 1183-1197, 1184-1198, 1185-1199, 1186-1200,
1187-1201, 1188-1202, 1189-1203, 1190-1204, 1191-1205, 1192-1206,
1193-1207, 1194-1208, 1195-1209, 1196-1210, 1197-1211, 1198-1212,
1199-1213, 1200-1214, 1201-1215, 1202-1216, 1203-1217, 1204-1218,
1205-1219, 1206-1220, 1207-1221, 1208-1222, 1209-1223, 1210-1224,
1211-1225, 1212-1226, 1213-1227, 1214-1228, 1215-1229, 1216-1230,
1217-1231, 1218-1232, 1219-1233, 1220-1234, 1221-1235, 1222-1236,
1223-1237, 1224-1238, 1225-1239, 1226-1240, 1227-1241, 1228-1242,
1229-1243, 1230-1244, 1231-1245, 1232-1246, 1233-1247, 1234-1248,
1235-1249, 1236-1250, 1237-1251, 1238-1252, 1239-1253, 1240-1254,
1241-1255, 1242-1256, 1243-1257, 1244-1258, 1245-1259, 1246-1260,
1247-1261, 1248-1262, 1249-1263, 1250-1264, 1251-1265, 1252-1266,
1253-1267, 1254-1268, 1255-1269, 1256-1270, 1257-1271, 1258-1272,
1259-1273, 1260-1274, 1261-1275, 1262-1276, 1263-1277, 1264-1278,
1265-1279, 1266-1280, 1267-1281, 1268-1282, 1269-1283, 1270-1284,
1271-1285, 1272-1286, 1273-1287, 1274-1288, 1275-1289, 1276-1290,
1277-1291, 1278-1292, 1279-1293, 1280-1294, 1281-1295, 1282-1296,
1283-1297, 1284-1298, 1285-1299, 1286-1300, 1287-1301, 1288-1302,
1289-1303, 1290-1304, 1291-1305, 1292-1306, 1293-1307, 1294-1308,
1295-1309, 1296-1310, 1297-1311, 1298-1312, 1299-1313, 1300-1314,
1301-1315, 1302-1315, 1303-1317, 1304-1318, 1305-1319, 1306-1320,
1307-1321, 1308-1322, 1309-1323, 1310-1324, 1311-1325, 1312-1326,
1313-1327, 1314-1328, 1315-1329, 1316-1330, 1317-1331, 1318-1332,
1319-1333, 1320-1334, 1321-1335, 1322-1336, 1323-1337, 1324-1338,
1325-1339, 1326-1340, 1327-1341, 1328-1342, 1329-1343, 1330-1344,
1331-1345, 1332-1346, 1333-1347, 1334-1348, 1335-1349, 1336-1350,
1337-1351, 1338-1352, 1339-1353, 1340-1354, 1341-1355, 1342-1356,
1343-1357, 1344-1358, 1345-1359, 1346-1360, 1347-1361, 1348-1362,
1349-1363, 1350-1364, 1351-1365, 1352-1366, 1353-1367, 1354-1368,
1355-1369, 1356-1370, 1357-1371, 1358-1372, 1359-1373, 1360-1374,
1361-1375, 1362-1376, 1363-1377, 1364-1378, 1365-1379, 1366-1380,
1367-1381, 1368-1382, 1369-1383, 1370-1384, 1371-1385, 1372-1386,
1373-1387, 1374-1388, 1375-1389, 1376-1390, 1377-1391, 1378-1392,
1379-1393, 1380-1394, 1381-1395, 1382-1396, 1383-1397, 1384-1398,
1385-1399, 1386-1400, 1387-1401, 1388-1402, 1389-1403, 1390-1404,
1391-1405, 1392-1406, 1393-1407, 1394-1408, 1395-1409, 1396-1410,
1397-1411, 1398-1412, 1399-1413, 1400-1414, 1401-1415, 1402-1416,
1403-1417, 1404-1418, 1405-1419, 1406-1420, 1407-1421, 1408-1422,
1409-1423, 1410-1424, 1411-1425, 1412-1426, 1413-1427, 1414-1428,
1415-1429, 1416-1430, 1417-1431, 1418-1432, 1419-1433, 1420-1434,
1421-1435, 1422-1436, 1423-1437, 1424-1438, 1425-1439, 1426-1440,
1427-1441, 1428-1442, 1429-1443, 1430-1444, 1431-1445, 1432-1446,
1433-1447, 1434-1448, 1435-1449, 1436-1450, 1437-1451, 1438-1452,
1439-1453, 1440-1454, 1441-1455, 1442-1456, 1443-1457, 1444-1458,
1445-1459, 1446-1460, 1447-1461, 1448-1462, 1449-1463, 1450-1464,
1451-1465, 1452-1466, 1453-1467, 1454-1468, 1455-1469, 1456-1470,
1457-1471, 1458-1472, 1459-1473, 1460-1474, 1461-1475, 1462-1476,
1463-1477, 1464-1478, 1465-1479, 1466-1480, 1467-1481, 1468-1482,
1469-1483, 1470-1484, 1471-1485, 1472-1486, 1473-1487, 1474-1488,
1475-1489, 1476-1490, 1477-1491, 1478-1492, 1479-1493, 1480-1494,
1481-1495, 1482-1496, 1483-1497, 1484-1498, 1485-1499, 1486-1500,
1487-1501, 1488-1502, 1489-1503, 1490-1504, 1491-1505, 1492-1506,
1493-1507, 1494-1508, 1495-1509, 1496-1510, 1497-1511, 1498-1513,
1499-1514, 1500-1515, 1501-1515, 1502-1516, 1503-1517, 1504-1518,
1505-1519, 1506-1520, 1507-1521, 1508-1522, 1509-1523, 1510-1524,
1511-1525, 1512-1526, 1513-1527, 1514-1528, 1515-1529, 1516-1530,
1517-1531, 1518-1532, 1519-1533, 1520-1534, 1521-1535, 1522-1536,
1523-1537, 1524-1538, 1525-1539, 1526-1540, 1527-1541, 1528-1542,
1529-1543, 1530-1544, 1531-1545, 1532-1546, 1533-1547, 1534-1548,
1535-1549, 1536-1550, 1537-1551, 1538-1552, 1539-1553, 1540-1554,
1541-1555, 1542-1556, 1543-1557, 1544-1558, 1545-1559, 1546-1560,
1547-1561, 1548-1562, 1549-1563, 1550-1564, 1551-1565, 1552-1566,
1553-1567, 1554-1568, 1555-1569, 1556-1570, 1557-1571, 1558-1572,
1559-1573, 1560-1574, 1561-1575, 1562-1576, 1563-1577, 1564-1578,
1565-1579, 1566-1580, 1567-1581, 1568-1582, 1569-1583, 1570-1584,
1571-1585, 1572-1586, 1573-1587, 1574-1588, 1575-1589, 1576-1590,
1577-1591, 1578-1592, 1579-1593, 1580-1594, 1581-1595, 1582-1596,
1583-1597, 1584-1598, 1585-1599, 1586-1600, 1587-1601, 1588-1602,
1589-1603, 1590-1604, 1591-1605, 1592-1606, 1593-1607, 1594-1608,
1595-1609, 1596-1610, 1597-1611, 1598-1612, 1599-1613, 1600-1614,
1601-1615, 1602-1616, 1603-1617, 1604-1618, 1605-1619, 1606-1620,
1607-1621, 1608-1622, 1609-1623, 1610-1624, 1611-1625, 1612-1626,
1613-1627, 1614-1628, 1615-1629, 1616-1630, 1617-1631, 1618-1632,
1619-1633, 1620-1634, 1621-1635, 1622-1636, 1623-1637, 1624-1638,
1625-1639, 1626-1640, 1627-1641, 1628-1642, 1629-1643, 1630-1644,
1631-1645, 1632-1646, 1633-1647, 1634-1648, 1635-1649, 1636-1650,
1637-1651, 1638-1652, 1639-1653, 1640-1654, 1641-1655, 1642-1656,
1643-1657, 1644-1658, 1645-1659, 1646-1660, 1647-1661, 1648-1662,
1649-1663, 1650-1664, 1651-1665, 1652-1666, 1653-1667, 1654-1668,
1655-1669, 1656-1670, 1657-1671, 1658-1672, 1659-1673, 1660-1674,
1661-1675, 1662-1676, 1663-1677, 1664-1678, 1665-1679, 1666-1680,
1667-1681, 1668-1682, 1669-1683, 1670-1684, 1671-1685, 1672-1686,
1673-1687, 1674-1688, 1675-1689, 1676-1690, 1677-1691, 1678-1692,
1679-1693, 1680-1694, 1681-1695, 1682-1696, 1683-1697, 1684-1698,
1685-1699, 1686-1700, 1687-1701, 1688-1702, 1689-1703, 1690-1704,
1691-1705, 1692-1706, 1693-1707, 1694-1708, 1695-1709, 1696-1710,
1697-1711, 1698-1712, 1699-1713, 1700-1714, 1701-1715, 1702-1716,
1703-1717, 1704-1718, 1705-1719, 1706-1720, 1707-1721, 1708-1722,
1709-1723, 1710-1724, 1711-1725, 1712-1726, 1713-1727, 1714-1728,
1715-1729, 1716-1730, 1717-1731, 1718-1732, 1719-1733, 1720-1734,
1721-1735, 1722-1736, 1723-1737, 1724-1738, 1725-1739, 1726-1740,
1727-1741, 1728-1742, 1729-1743, 1730-1744, 1731-1745, 1732-1746,
1733-1747, 1734-1748, 1735-1749, 1736-1750, 1737-1751, 1738-1752,
1739-1753, 1740-1754, 1741-1755, 1742-1756, 1743-1757, 1744-1758,
1745-1759, 1746-1760, 1747-1761, 1748-1762, 1749-1763, 1750-1764,
1751-1765, 1752-1766, 1753-1767,
1754-1768, 1755-1769, 1756-1770, 1757-1771, 1758-1772, 1759-1773,
1760-1774, 1761-1775, 1762-1776, 1763-1777, 1764-1778, 1765-1779,
1766-1780, 1767-1781, 1768-1782, 1769-1783, 1770-1784, 1771-1785,
1772-1786, 1773-1787, 1774-1788, 1775-1789, 1776-1790, 1777-1791,
1778-1792, 1779-1793, 1780-1794, 1781-1795, 1782-1796, 1783-1797,
1784-1798, 1785-1799, 1786-1800, 1787-1801, 1788-1802, 1789-1803,
1790-1804, 1791-1805, 1792-1806, 1793-1807, 1794-1808, 1795-1809,
1796-1810, 1797-1811, 1798-1812, 1799-1813, 1800-1814, 1801-1815,
1802-1816, 1803-1817, 1804-1818, 1805-1819, 1806-1820, 1807-1821,
1808-1822, 1809-1823, 1810-1824, 1811-1825, 1812-1826, 1813-1827,
1814-1828, 1815-1829, 1816-1830, 1817-1831, 1818-1832, 1819-1833,
1820-1834, 1821-1835, 1822-1836, 1823-1837, 1824-1838, 1825-1839,
1826-1840, 1827-1841, 1828-1842, 1829-1843, 1830-1844, 1831-1845,
1832-1846, 1833-1847, 1834-1848, 1835-1849, 1836-1850, 1837-1851,
1838-1852, 1839-1853, 1840-1854, 1841-1855, 1842-1856, 1843-1857,
1844-1858, 1845-1859, 1846-1860, 1847-1861, 1848-1862, 1849-1863,
1850-1864, 1851-1865, 1852-1866, 1853-1867, 1854-1868, 1855-1869,
1856-1870, 1857-1871, 1858-1872, 1859-1873, 1860-1874, 1861-1875,
1862-1876, 1863-1877, 1864-1878, 1865-1879, 1866-1880, 1867-1881,
1868-1882, 1869-1883, 1870-1884, 1871-1885, 1872-1886, 1873-1887,
1874-1888, 1875-1889, 1876-1890, 1877-1891, 1878-1892, 1879-1893,
1880-1894, 1881-1895, 1882-1896, 1883-1897, 1884-1898, 1885-1899,
1886-1900, 1887-1901, 1888-1902, 1889-1903, 1890-1904, 1891-1905,
1892-1906, 1893-1907, 1894-1908, 1895-1909, 1896-1910, 1897-1911,
1898-1912, 1899-1913, 1900-1914, 1901-1915, 1902-1916, 1903-1917,
1904-1918, 1905-1919, 1906-1920, 1907-1921, 1908-1922, 1909-1923,
1910-1924, 1911-1925, 1912-1926, 1913-1927, 1914-1928, 1915-1929,
1916-1930, 1917-1931, 1918-1932, 1919-1933, 1920-1934, 1921-1935,
1922-1936, 1923-1937, 1924-1938, 1925-1939, 1926-1940, 1927-1941,
1928-1942, 1929-1943, 1930-1944, 1931-1945, 1932-1946, 1933-1947,
1934-1948, 1935-1949, 1936-1950, 1937-1951, 1938-1952, 1939-1953,
1940-1954, 1941-1955, 1942-1956, 1943-1957, 1944-1958, 1945-1959,
1946-1960, 1947-1961, 1948-1962, 1949-1963, 1950-1964, 1951-1965,
1952-1966, 1953-1967, 1954-1968, 1955-1969, 1956-1970, 1957-1971,
1958-1972, 1959-1973, 1960-1974, 1961-1975, 1962-1976, 1963-1977,
1964-1978, 1965-1979, 1966-1980, 1967-1981, 1968-1982, 1969-1983,
1970-1984, 1971-1985, 1972-1986, 1973-1987, 1974-1988, 1975-1989,
1976-1990, 1977-1991, 1978-1992, 1979-1993, 1980-1994, 1981-1995,
1982-1996, 1983-1997, 1984-1998, 1985-1999, 1986-2000, 1987-2001,
1988-2002, 1989-2003, 1990-2004, 1991-2005, 1992-2006, 1993-2007,
1994-2008, 1995-2009, 1996-2010, 1997-2011, 1998-2012, 1999-2013,
2000-2014, 2001-2015, 2002-2016, 2003-2017, 2004-2018, 2005-2019,
2006-2020, 2007-2021, 2008-2022, 2009-2023, 2010-2024, 2011-2025,
2012-2026, 2013-2027, 2014-2028, 2015-2029, 2016-2030, 2017-2031,
2018-2032, 2019-2033, 2020-2034, 2021-2035, 2022-2036, 2023-2037,
2024-2038, 2025-2039, 2026-2040, 2027-2041, 2028-2042, 2029-2043,
2030-2044, 2031-2045, 2032-2046, 2033-2047, 2034-2048, 2035-2049,
2036-2050, 2037-2051, 2038-2052, 2039-2053, 2040-2054, 2041-2055,
2042-2056, 2043-2057, 2044-2058, 2045-2059, 2046-2060, 2047-2061,
2048-2062, 2049-2063, 2050-2064, 2051-2065, 2052-2066, 2053-2067,
2054-2068, 2055-2069, 2056-2070, 2057-2071, 2058-2072, 2059-2073,
2060-2074, 2061-2075, 2062-2076, 2063-2077, 2064-2078, 2065-2079,
2066-2080, 2067-2081, 2068-2082, 2069-2083, 2070-2084, 2071-2085,
2072-2086, 2073-2087, 2074-2088, 2075-2089, 2076-2090, 2077-2091,
2078-2092, 2079-2093, 2080-2094, 2081-2095, 2082-2096, 2083-2097,
2084-2098, 2085-2099, 2086-2100, 2087-2101, 2088-2102, 2089-2103,
2090-2104, 2091-2105, 2092-2106, 2093-2107, 2094-2108, 2095-2109,
2096-2110, 2097-2111, 2098-2112, 2099-2113, 2100-2114, 2101-2115,
2102-2116, 2103-2117, 2104-2118, 2105-2119, 2106-2120, 2107-2121,
2108-2122, 2109-2123, 2110-2124, 2111-2125, 2112-2126, 2113-2127,
2114-2128, 2115-2129, 2116-2130, 2117-2131, 2118-2132, 2119-2133,
2120-2134, 2121-2135, 2122-2136, 2123-2137, 2124-2138, 2125-2139,
2126-2140, 2127-2141, 2128-2142, 2129-2143, 2130-2144, 2131-2145,
2132-2146, 2133-2147, 2134-2148, 2135-2149, 2136-2150, 2137-2151,
2138-2152, 2139-2153, 2140-2154, 2141-2155, 2142-2156, 2143-2157,
2144-2158, 2145-2159, 2146-2160, 2147-2161, 2148-2162, 2149-2163,
2150-2164, 2151-2165, 2152-2166, 2153-2167, 2154-2168, 2155-2169,
2156-2170, 2157-2171, 2158-2172, 2159-2173, 2160-2174, 2161-2175,
2162-2176, 2163-2177, 2164-2178, 2165-2179, 2166-2180, 2167-2181,
2168-2182, 2169-2183, 2170-2184, 2171-2185, 2172-2186, 2173-2187,
2174-2188, 2175-2189, 2176-2190, 2177-2191, 2178-2192, 2179-2193,
2180-2194, 2181-2195, 2182-2196, 2183-2197, 2184-2198, 2185-2199,
2186-2200, 2187-2201, 2188-2202, 2189-2203, 2190-2204, 2191-2205,
2192-2206, 2193-2207, 2194-2208, 2195-2209, 2196-2210, 2197-2211,
2198-2212, 2199-2213, 2200-2214, 2201-2215, 2202-2216, 2203-2217,
2204-2218, 2205-2219, 2206-2220, 2207-2221, 2208-2222, 2209-2223,
2210-2224, 2211-2225, 2212-2226, 2213-2227, 2214-2228, 2215-2229,
2216-2230, 2217-2231, 2218-2232, 2219-2233, 2220-2234, 2221-2235,
2222-2236, 2223-2237, 2224-2238, 2225-2239, 2226-2240, 2227-2241,
2228-2242, 2229-2243, 2230-2244, 2231-2245, 2232-2246, 2233-2247,
2234-2248, 2235-2249, 2236-2250, 2237-2251, 2238-2252, 2239-2253,
2240-2254, 2241-2255, 2242-2256, 2243-2257, 2244-2258, 2245-2259,
2246-2260, 2247-2261, 2248-2262, 2249-2263, 2250-2264, 2251-2265,
2252-2266, 2253-2267, 2254-2268, 2255-2269, 2256-2270, 2257-2271,
2258-2272, 2259-2273, 2260-2274, 2261-2275, 2262-2276, 2263-2277,
2264-2278, 2265-2279, 2266-2280, 2267-2281, 2268-2282, 2269-2283,
2270-2284, 2271-2285, 2272-2286, 2273-2287, 2274-2288, 2275-2289,
2276-2290, 2277-2291, 2278-2292, 2279-2293, 2280-2294, 2281-2295,
2282-2296, 2283-2297, 2284-2298, 2285-2299, 2286-2300, 2287-2301,
2288-2302, 2289-2303, 2290-2304, 2291-2305, 2292-2306, 2293-2307,
2294-2308, 2295-2309, 2296-2310, 2297-2311, 2298-2312, 2299-2313,
2300-2314, 2301-2315, 2302-2315, 2303-2317, 2304-2318, 2305-2319,
2306-2320, 2307-2321, 2308-2322, 2309-2323, 2310-2324, 2311-2325,
2312-2326, 2313-2327, 2314-2328, 2315-2329, 2316-2330, 2317-2331,
2318-2332, 2319-2333, 2320-2334, 2321-2335, 2322-2336, 2323-2337,
2324-2338, 2325-2339, 2326-2340, 2327-2341, 2328-2342, 2329-2343,
2330-2344, 2331-2345, 2332-2346, 2333-2347, 2334-2348, 2335-2349,
2336-2350, 2337-2351, 2338-2352, 2339-2353, 2340-2354, 2341-2355,
2342-2356, 2343-2357, 2344-2358, 2345-2359, 2346-2360, 2347-2361,
2348-2362, 2349-2363, 2350-2364, 2351-2365, 2352-2366, 2353-2367,
2354-2368, 2355-2369, 2356-2370, 2357-2371, 2358-2372, 2359-2373,
2360-2374, 2361-2375, 2362-2376, 2363-2377, 2364-2378, 2365-2379,
2366-2380, 2367-2381, 2368-2382, 2369-2383, 2370-2384, 2371-2385,
2372-2386, 2373-2387, 2374-2388, 2375-2389, 2376-2390, 2377-2391,
2378-2392, 2379-2393, 2380-2394, 2381-2395, 2382-2396, 2383-2397,
2384-2398, 2385-2399, 2386-2400, 2387-2401, 2388-2402, 2389-2403,
2390-2404, 2391-2405, 2392-2406, 2393-2407, 2394-2408, 2395-2409,
2396-2410, 2397-2411, 2398-2412, 2399-2413, 2400-1412, 2401-2415,
2402-2416, 2403-2417, 2404-2418, 2405-2419, 2406-2420, 2407-2421,
2408-2422, 2409-2423, 2410-2424, 2411-2425, 2412-2426, 2413-2427,
2414-2428, 2415-2429, 2416-2430, 2417-2431, 2418-2432, 2419-2433,
2420-2434, 2421-2435, 2422-2436, 2423-2437, 2424-2438, 2425-2439,
2426-2440, 2427-2441, 2428-2442, 2429-2443, 2430-2444, 2431-2445,
2432-2446, 2433-2447, 2434-2448, 2435-2449, 2436-2450, 2437-2451,
2438-2452, 2439-2453, 2440-2454, 2441-2455, 2442-2456, 2443-2457,
2444-2458, 2445-2459, 2446-2460, 2447-2461, 2448-2462, 2449-2463,
2450-2464, 2451-2465, 2452-2466, 2453-2467, 2454-2468, 2455-2469,
2456-2470, 2457-2471, 2458-2472, 2459-2473, and 2460-2474 of SEQ ID
NO:2.
EXAMPLE 8
ANTISENSE OLIGONUCLEOTIDES OF IGF-I RECEPTOR
Antisense oligonucleotides to the IGF-I receptor may be selected
from molecules capable of interacting with one or more
oligonucleotides selected from oligonucleotides having the sequence
of nucleotides 1-15, 2-16, 3-17, 4-18, 5-19, 6-20, 7-21, 8-22,
9-23, 10-24, 11-25, 12-26, 13-27, 14-28, 15-29, 16-30, 17-31,
18-32, 19-33, 20-34, 21-35, 22-36, 23-37, 24-38, 25-39, 26-40,
27-41, 28-42, 29-43, 30-44, 31-45, 32-46, 33-47, 34-48, 35-49,
36-50, 37-51, 38-52, 39-53, 40-54, 41-55, 42-56, 43-57, 44-58,
45-59, 46-60, 47-61, 48-62, 49-63, 50-64, 51-65, 52-66, 53-67,
54-68, 55-69, 56-70, 57-71, 58-72, 59-73, 60-74, 61-75, 62-76,
63-77, 64-78, 65-79, 66-80, 67-81, 68-82, 69-83, 70-84, 71-85,
72-86, 73-87, 74-88, 75-89, 76-90, 77-91, 78-92, 79-93, 80-94,
81-95, 82-96, 83-97, 84-98, 85-99, 86-100, 87-101, 88-102, 89-103,
90-104, 91-105, 92-106, 93-107, 94-108, 95-109, 96-110, 97-111,
98-112, 99-113, 100-114, 101-115, 102-116, 103-117, 104-118,
105-119, 106-120, 107-121, 108-122, 109-123, 110-124, 111-125,
112-126, 113-127, 114-128, 115-129, 116-130, 117-131, 118-132,
119-133, 120-134, 121-135, 122-136, 123-137, 124-138, 125-139,
126-140, 127-141, 128-142, 129-143, 130-144, 131-145, 132-146,
133-147, 134-148, 135-149, 136-150, 137-151, 138-152, 139-153,
140-154, 141-155, 142-156, 143-157, 144-158, 145-159, 146-160,
147-161, 148-162, 149-163, 150-164, 151-165, 152-166, 153-167,
154-168, 155-169, 156-170, 157-171, 158-172, 159-173, 160-174,
161-175, 162-176, 163-177, 164-178, 165-179, 166-180, 167-181,
168-182, 169-183, 170-184, 171-185, 172-186, 173-187, 174-188,
175-189, 176-190, 177-191, 178-192, 179-193, 180-194, 181-195,
182-196, 183-197, 184-198, 185-199, 186-200, 187-201, 188-202,
189-203, 190-204, 191-205, 192-206, 193-207, 194-208, 195-209,
196-210, 197-211, 198-212, 199-213, 200-214, 201-215, 202-216,
203-217, 204-218, 205-219, 206-220, 207-221, 208-222, 209-223,
210-224, 211-225, 212-226, 213-227, 214-228, 215-229, 216-230,
217-231, 218-232, 219-233, 220-234, 221-235, 222-236, 223-237,
224-238, 225-239, 226-240, 227-241, 228-242, 229-243, 230-244,
231-245, 232-246, 233-247, 234-248, 235-249, 236-250, 237-251,
238-252, 239-253, 240-254, 241-255, 242-256, 243-257, 244-258,
245-259, 246-260, 247-261, 248-262, 249-263, 250-264, 251-265,
252-266, 253-267, 254-268, 255-269, 256-270, 257-271, 258-272,
259-273, 260-274, 261-275, 262-276, 263-277, 264-278, 265-279,
266-280, 267-281, 268-282, 269-283, 270-284, 271-285, 272-286,
273-287, 274-288, 275-289, 276-290, 277-291, 278-292, 279-293,
280-294, 281-295, 282-296, 283-297, 284-298, 285-299, 286-300,
287-301, 288-302, 289-303, 290-304, 291-305, 292-306, 293-307,
294-308, 295-309, 296-310, 297-311, 298-312, 299-313, 300-314,
301-315, 302-315, 303-317, 304-318, 305-319, 306-320, 307-321,
308-322, 309-323, 310-324, 311-325, 312-326, 313-327, 314-328,
315-329, 316-330, 317-331, 318-332, 319-333, 320-334, 321-335,
322-336, 323-337, 324-338, 325-339, 326-340, 327-341, 328-342,
329-343, 330-344, 331-345, 332-346, 333-347, 334-348, 335-349,
336-350, 337-351, 338-352, 339-353, 340-354, 341-355, 342-356,
343-357, 344-358, 345-359, 346-360, 347-361, 348-362, 349-363,
350-364, 351-365, 352-366, 353-367, 354-368, 355-369, 356-370,
357-371, 358-372, 359-373, 360-374, 361-375, 362-376, 363-377,
364-378, 365-379, 366-380, 367-381, 368-382, 369-383, 370-384,
371-385, 372-386, 373-387, 374-388, 375-389, 376-390, 377-391,
378-392, 379-393, 380-394, 381-395, 382-396, 383-397, 384-398,
385-399, 386-400, 387-401, 388-402, 389-403, 390-404, 391-405,
392-406, 393-407, 394-408, 395-409, 396-410, 397-411, 398-412,
399-413, 400-414, 401-415, 402-416, 403-417, 404-418, 405-419,
406-420, 407-421, 408-422, 409-423, 410-424, 411-425, 412-426,
413-427, 414-428, 415-429, 416-430, 417-431, 418-432, 419-433,
420-434, 421-435, 422-436, 423-437, 424-438, 425-439, 426-440,
427-441, 428-442, 429-443, 430-444, 431-445, 432-446, 433-447,
434-448, 435-449, 436-450, 437-451, 438-452, 439-453, 440-454,
441-455, 442-456, 443-457, 444-458, 445-459, 446-460, 447-461,
448-462, 449-463, 450-464, 451-465, 452-466, 453-467, 454-468,
455-469, 456-470, 457-471, 458-472, 459-473, 460-474, 461-475,
462-476, 463-477, 464-478, 465-479, 466-480, 467-481, 468-482,
469-483, 470-484, 471-485, 472-486, 473-487, 474-488, 475-489,
476-490, 477-491, 478-492, 479-493, 480-494, 481-495, 482-496,
483-497, 484-498, 485-499, 486-500, 487-501, 488-502, 489-503,
490-504, 491-505, 492-506, 493-507, 494-508, 495-509, 496-510,
497-511, 498-513, 499-514, 500-515, 501-515, 502-516, 503-517,
504-518, 505-519, 506-520, 507-521, 508-522, 509-523, 510-524,
511-525, 512-526, 513-527, 514-528, 515-529, 516-530, 517-531,
518-532, 519-533, 520-534, 521-535, 522-536, 523-537, 524-538,
525-539, 526-540, 527-541, 528-542, 529-543, 530-544, 531-545,
532-546, 533-547, 534-548, 535-549, 536-550, 537-551, 538-552,
539-553, 540-554, 541-555, 542-556, 543-557, 544-558, 545-559,
546-560, 547-561, 548-562, 549-563, 550-564, 551-565, 552-566,
553-567, 554-568, 555-569, 556-570, 557-571, 558-572, 559-573,
560-574, 561-575, 562-576, 563-577, 564-578, 565-579, 566-580,
567-581, 568-582, 569-583, 570-584, 571-585, 572-586, 573-587,
574-588, 575-589, 576-590, 577-591, 578-592, 579-593, 580-594,
581-595, 582-596, 583-597, 584-598, 585-599, 586-600, 587-601,
588-602, 589-603, 590-604, 591-605, 592-606, 593-607, 594-608,
595-609, 596-610, 597-611, 598-612, 599-613, 600-614, 601-615,
602-616, 603-617, 604-618, 605-619, 606-620, 607-621, 608-622,
609-623, 610-624, 611-625, 612-626, 613-627, 614-628, 615-629,
616-630, 617-631, 618-632, 619-633, 620-634, 621-635, 622-636,
623-637, 624-638, 625-639, 626-640, 627-641, 628-642, 629-643,
630-644, 631-645, 632-646, 633-647, 634-648, 635-649, 636-650,
637-651, 638-652, 639-653, 640-654, 641-655, 642-656, 643-657,
644-658, 645-659, 646-660, 647-661, 648-662, 649-663, 650-664,
651-665, 652-666, 653-667, 654-668, 655-669, 656-670, 657-671,
658-672, 659-673, 660-674, 661-675, 662-676, 663-677, 664-678,
665-679, 666-680, 667-681, 668-682, 669-683, 670-684, 671-685,
672-686, 673-687, 674-688, 675-689, 676-690, 677-691, 678-692,
679-693, 680-694, 681-695, 682-696, 683-697, 684-698, 685-699,
686-700, 687-701, 688-702, 689-703, 690-704, 691-705, 692-706,
693-707, 694-708, 695-709, 696-710, 697-711, 698-712, 699-713,
700-714, 701-715, 702-716, 703-717, 704-718, 705-719, 706-720,
707-721, 708-722, 709-723, 710-724, 711-725, 712-726, 713-727,
714-728, 715-729, 716-730, 717-731, 718-732, 719-733, 720-734,
721-735, 722-736, 723-737, 724-738, 725-739, 726-740, 727-741,
728-742, 729-743, 730-744, 731-745, 732-746, 733-747, 734-748,
735-749, 736-750, 737-751, 738-752, 739-753, 740-754, 741-755,
742-756, 743-757, 744-758, 745-759, 746-760, 747-761, 748-762,
749-763, 750-764, 751-765, 752-766, 753-767, 754-768, 755-769,
756-770, 757-771, 758-772, 759-773, 760-774, 761-775, 762-776,
763-777, 764-778, 765-779, 766-780, 767-781, 768-782, 769-783,
770-784, 771-785, 772-786, 773-787, 774-788, 775-789, 776-790,
777-791, 778-792, 779-793, 780-794, 781-795, 782-796, 783-797,
784-798, 785-799, 786-800, 787-801, 788-802, 789-803, 790-804,
791-805, 792-806, 793-807, 794-808, 795-809, 796-810, 797-811,
798-812, 799-813, 800-814, 801-815, 802-816, 803-817, 804-818,
805-819, 806-820, 807-821, 808-822, 809-823, 810-824, 811-825,
812-826, 813-827, 814-828, 815-829, 816-830, 817-831, 818-832,
819-833, 820-834, 821-835, 822-836, 823-837, 824-838, 825-839,
826-840, 827-841, 828-842, 829-843, 830-844, 831-845, 832-846,
833-847, 834-848, 835-849, 836-850, 837-851, 838-852, 839-853,
840-854, 841-855, 842-856, 843-857, 844-858, 845-859, 846-860,
847-861, 848-862, 849-863, 850-864, 851-865, 852-866, 853-867,
854-868, 855-869, 856-870, 857-871, 858-872, 859-873, 860-874,
861-875, 862-876, 863-877, 864-878, 865-879, 866-880, 867-881,
868-882, 869-883, 870-884, 871-885, 872-886, 873-887, 874-888,
875-889, 876-890, 877-891, 878-892, 879-893, 880-894, 881-895,
882-896, 883-897, 884-898, 885-899, 886-900, 887-901, 888-902,
889-903, 890-904, 891-905, 892-906, 893-907, 894-908, 895-909,
896-910, 897-911, 898-912, 899-913, 900-914, 901-915, 902-916,
903-917, 904-918, 905-919, 906-920, 907-921, 908-922, 909-923,
910-924, 911-925, 912-926, 913-927, 914-928, 915-929, 916-930,
917-931, 918-932, 919-933, 920-934, 921-935, 922-936, 923-937,
924-938, 925-939, 926-940, 927-941, 928-942, 929-943, 930-944,
931-945, 932-946, 933-947, 934-948, 935-949, 936-950, 937-951,
938-952, 939-953, 940-954, 941-955, 942-956, 943-957, 944-958,
945-959, 946-960, 947-961, 948-962, 949-963, 950-964, 951-965,
952-966, 953-967, 954-968, 955-969, 956-970, 957-971, 958-972,
959-973, 960-974, 961-975, 962-976, 963-977, 964-978, 965-979,
966-980, 967-981, 968-982, 969-983, 970-984, 971-985, 972-986,
973-987, 974-988, 975-989, 976-990, 977-991, 978-992, 979-993,
980-994, 981-995, 982-996, 983-997, 984-998, 985-999, 986-1000,
987-1001, 988-1002, 989-1003, 990-1004, 991-1005, 992-1006,
993-1007, 994-1008, 995-1009, 996-1010, 997-1011, 998-1012,
999-1013, 1000-1014, 1001-1015, 1002-1016, 1003-1017, 1004-1018,
1005-1019, 1006-1020, 1007-1021, 1008-1022, 1009-1023, 1010-1024,
1011-1025, 1012-1026, 1013-1027, 1014-1028, 1015-1029, 1016-1030,
1017-1031, 1018-1032, 1019-1033, 1020-1034, 1021-1035, 1022-1036,
1023-1037, 1024-1038, 1025-1039, 1026-1040, 1027-1041, 1028-1042,
1029-1043, 1030-1044, 1031-1045, 1032-1046, 1033-1047, 1034-1048,
1035-1049, 1036-1050, 1037-1051, 1038-1052, 1039-1053, 1040-1054,
1041-1055, 1042-1056, 1043-1057, 1044-1058, 1045-1059, 1046-1060,
1047-1061, 1048-1062, 1049-1063, 1050-1064, 1051-1065, 1052-1066,
1053-1067, 1054-1068, 1055-1069, 1056-1070, 1057-1071, 1058-1072,
1059-1073, 1060-1074, 1061-1075, 1062-1076, 1063-1077, 1064-1078,
1065-1079, 1066-1080, 1067-1081, 1068-1082, 1069-1083, 1070-1084,
1071-1085, 1072-1086, 1073-1087, 1074-1088, 1075-1089, 1076-1090,
1077-1091, 1078-1092, 1079-1093, 1080-1094, 1081-1095, 1082-1096,
1083-1097, 1084-1098, 1085-1099, 1086-1100, 1087-1101, 1088-1102,
1089-1103, 1090-1104, 1091-1105, 1092-1106, 1093-1107, 1094-1108,
1095-1109, 1096-1110, 1097-1111, 1098-1112, 1099-1113, 1100-1114,
1101-1115, 1102-1116, 1103-1117, 1104-1118, 1105-1119, 1106-1120,
1107-1121, 1108-1122, 1109-1123, 1110-1124, 1111-1125, 1112-1126,
1113-1127, 1114-1128, 1115-1129, 1116-1130, 1117-1131, 1118-1132,
1119-1133, 1120-1134, 1121-1135, 1122-1136, 1123-1137, 1124-1138,
1125-1139, 1126-1140, 1127-1141, 1128-1142, 1129-1143, 1130-1144,
1131-1145, 1132-1146, 1133-1147, 1134-1148, 1135-1149, 1136-1150,
1137-1151, 1138-1152, 1139-1153, 1140-1154, 1141-1155, 1142-1156,
1143-1157, 1144-1158, 1145-1159, 1146-1160, 1147-1161, 1148-1162,
1149-1163, 1150-1164, 1151-1165, 1152-1166, 1153-1167, 1154-1168,
1155-1169, 1156-1170, 1157-1171, 1158-1172, 1159-1173, 1160-1174,
1161-1175, 1162-1176, 1163-1177, 1164-1178, 1165-1179, 1166-1180,
1167-1181, 1168-1182, 1169-1183, 1170-1184, 1171-1185, 1172-1186,
1173-1187, 1174-1188, 1175-1189, 1176-1190, 1177-1191, 1178-1192,
1179-1193, 1180-1194, 1181-1195, 1182-1196, 1183-1197, 1184-1198,
1185-1199, 1186-1200, 1187-1201, 1188-1202, 1189-1203, 1190-1204,
1191-1205, 1192-1206, 1193-1207, 1194-1208, 1195-1209, 1196-1210,
1197-1211, 1198-1212, 1199-1213, 1200-1214, 1201-1215, 1202-1216,
1203-1217, 1204-1218, 1205-1219, 1206-1220, 1207-1221, 1208-1222,
1209-1223, 1210-1224, 1211-1225, 1212-1226, 1213-1227, 1214-1228,
1215-1229, 1216-1230, 1217-1231, 1218-1232, 1219-1233, 1220-1234,
1221-1235, 1222-1236, 1223-1237, 1224-1238, 1225-1239, 1226-1240,
1227-1241, 1228-1242, 1229-1243, 1230-1244, 1231-1245, 1232-1246,
1233-1247, 1234-1248, 1235-1249, 1236-1250, 1237-1251, 1238-1252,
1239-1253, 1240-1254, 1241-1255, 1242-1256, 1243-1257, 1244-1258,
1245-1259, 1246-1260, 1247-1261, 1248-1262, 1249-1263, 1250-1264,
1251-1265, 1252-1266, 1253-1267, 1254-1268, 1255-1269, 1256-1270,
1257-1271, 1258-1272, 1259-1273, 1260-1274, 1261-1275, 1262-1276,
1263-1277, 1264-1278, 1265-1279, 1266-1280, 1267-1281, 1268-1282,
1269-1283, 1270-1284, 1271-1285, 1272-1286, 1273-1287, 1274-1288,
1275-1289, 1276-1290, 1277-1291, 1278-1292, 1279-1293, 1280-1294,
1281-1295, 1282-1296, 1283-1297, 1284-1298, 1285-1299, 1286-1300,
1287-1301, 1288-1302, 1289-1303, 1290-1304, 1291-1305, 1292-1306,
1293-1307, 1294-1308, 1295-1309, 1296-1310, 1297-1311, 1298-1312,
1299-1313, 1300-1314, 1301-1315, 1302-1317, 1303-1317, 1304-1318,
1305-1319, 1306-1320, 1307-1321, 1308-1322, 1309-1323, 1310-1324,
1311-1325, 1312-1326, 1313-1327, 1314-1328, 1315-1329, 1316-1330,
1317-1331, 1318-1332, 1319-1333, 1320-1334, 1321-1335, 1322-1336,
1323-1337, 1324-1338, 1325-1339, 1326-1340, 1327-1341, 1328-1342,
1329-1343, 1330-1344, 1331-1345, 1332-1346, 1333-1347, 1334-1348,
1335-1349, 1336-1350, 1337-1351, 1338-1352, 1339-1353, 1340-1354,
1341-1355, 1342-1356, 1343-1357, 1344-1358, 1345-1359, 1346-1360,
1347-1361, 1348-1362, 1349-1363, 1350-1364, 1351-1365, 1352-1366,
1353-1367, 1354-1368, 1355-1369, 1356-1370, 1357-1371, 1358-1372,
1359-1373, 1360-1374, 1361-1375, 1362-1376, 1363-1377, 1364-1378,
1365-1379, 1366-1380, 1367-1381, 1368-1382, 1369-1383, 1370-1384,
1371-1385, 1372-1386, 1373-1387, 1374-1388, 1375-1389, 1376-1390,
1377-1391, 1378-1392, 1379-1393, 1380-1394, 1381-1395, 1382-1396,
1383-1397, 1384-1398, 1385-1399, 1386-1400, 1387-1401, 1388-1402,
1389-1403, 1390-1404, 1391-1405, 1392-1406, 1393-1407, 1394-1408,
1395-1409, 1396-1410, 1397-1411, 1398-1412, 1399-1413, 1400-1414,
1401-1415, 1402-1416, 1403-1417, 1404-1418, 1405-1419, 1406-1420,
1407-1421, 1408-1422, 1409-1423, 1410-1424, 1411-1425, 1412-1426,
1413-1427, 1414-1428, 1415-1429, 1416-1430, 1417-1431, 1418-1432,
1419-1433, 1420-1434, 1421-1435, 1422-1436, 1423-1437, 1424-1438,
1425-1439, 1426-1440, 1427-1441, 1428-1442, 1429-1443, 1430-1444,
1431-1445, 1432-1446, 1433-1447, 1434-1448, 1435-1449, 1436-1450,
1437-1451, 1438-1452, 1439-1453, 1440-1454, 1441-1455, 1442-1456,
1443-1457, 1444-1458, 1445-1459, 1446-1460, 1447-1461, 1448-1462,
1449-1463, 1450-1464, 1451-1465, 1452-1466, 1453-1467, 1454-1468,
1455-1469, 1456-1470, 1457-1471, 1458-1472, 1459-1473, 1460-1474,
1461-1475, 1462-1476, 1463-1477, 1464-1478, 1465-1479, 1466-1480,
1467-1481, 1468-1482, 1469-1483, 1470-1484, 1471-1485, 1472-1486,
1473-1487, 1474-1488, 1475-1489, 1476-1490, 1477-1491, 1478-1492,
1479-1493, 1480-1494, 1481-1495, 1482-1496, 1483-1497, 1484-1498,
1485-1499, 1486-1500, 1487-1501, 1488-1502, 1489-1503, 1490-1504,
1491-1505, 1492-1506, 1493-1507, 1494-1508, 1495-1509, 1496-1510,
1497-1511, 1498-1513, 1499-1514, 1500-1515, 1501-1515, 1502-1516,
1503-1517, 1504-15183, 1505-1519, 1506-1520, 1507-1521, 1508-1522,
1509-1523, 1510-1524, 1511-1525, 1512-1526, 1513-1527, 1514-1528,
1515-1529, 1516-1530, 1517-1531, 1518-1532, 1519-1533, 1520-1534,
1521-1535, 1522-1536, 1523-1537, 1524-1538, 1525-1539, 1526-1540,
1527-1541, 1528-1542, 1529-1543, 1530-1544, 1531-1545, 1532-1546,
1533-1547, 1534-1548, 1535-1549, 1536-1550, 1537-1551, 1538-1552,
1539-1553, 1540-1554, 1541-1555, 1542-1556, 1543-1557, 1544-1558,
1545-1559, 1546-1560, 1547-1561, 1548-1562, 1549-1563, 1550-1564,
1551-1565, 1552-1566, 1553-1567, 1554-1568, 1555-1569, 1556-1570,
1557-1571, 1558-1572, 1559-1573, 1560-1574, 1561-1575, 1562-1576,
1563-1577, 1564-1578, 1565-1579, 1566-1580, 1567-1581, 1568-1582,
1569-1583, 1570-1584, 1571-1585, 1572-1586, 1573-1587, 1574-1588,
1575-1589, 1576-1590, 1577-1591, 1578-1592, 1579-1593, 1580-1594,
1581-1595, 1582-1596, 1583-1597, 1584-1598, 1585-1599, 1586-1600,
1587-1601, 1588-1602, 1589-1603, 1590-1604, 1591-1605, 1592-1606,
1593-1607, 1594-1608, 1595-1609, 1596-1610, 1597-1611, 1598-1612,
1599-1613, 1600-1614, 1601-1615, 1602-1616, 1603-1617, 1604-1618,
1605-1619, 1606-1620, 1607-1621, 1608-1622, 1609-1623, 1610-1624,
1611-1625, 1612-1626, 1613-1627, 1614-1628, 1615-1629, 1616-1630,
1617-1631, 1618-1632, 1619-1633, 1620-1634, 1621-1635, 1622-1636,
1623-1637, 1624-1638, 1625-1639, 1626-1640, 1627-1641, 1628-1642,
1629-1643, 1630-1644, 1631-1645, 1632-1646, 1633-1647, 1634-1648,
1635-1649, 1636-1650, 1637-1651, 1638-1652, 1639-1653, 1640-1654,
1641-1655, 1642-1656, 1643-1657, 1644-1658, 1645-1659, 1646-1660,
1647-1661, 1648-1662, 1649-1663, 1650-1664, 1651-1665, 1652-1666,
1653-1667, 1654-1668, 1655-1669, 1656-1670, 1657-1671, 1658-1672,
1659-1673, 1660-1674, 1661-1675, 1662-1676, 1663-1677, 1664-1678,
1665-1679, 1666-1680, 1667-1681, 1668-1682, 1669-1683, 1670-1684,
1671-1685, 1672-1686, 1673-1687, 1674-1688, 1675-1689, 1676-1690,
1677-1691, 1678-1692, 1679-1693, 1680-1694, 1681-1695, 1682-1696,
1683-1697, 1684-1698, 1685-1699, 1686-1700, 1687-1701, 1688-1702,
1689-1703, 1690-1704, 1691-1705, 1692-1706, 1693-1707, 1694-1708,
1695-1709, 1696-1710, 1697-1711, 1698-1712, 1699-1713, 1700-1714,
1701-1715, 1702-1716, 1703-1717, 1704-1718, 1705-1719, 1706-1720,
1707-1721, 1708-1722, 1709-1723, 1710-1724, 1711-1725, 1712-1726,
1713-1727, 1714-1728, 1715-1729, 1716-1730, 1717-1731, 1718-1732,
1719-1733, 1720-1734, 1721-1735, 1722-1736, 1723-1737, 1724-1738,
1725-1739, 1726-1740, 1727-1741, 1728-1742, 1729-1743, 1730-1744,
1731-1745, 1732-1746, 1733-1747, 1734-1748, 1735-1749, 1736-1750,
1737-1751, 1738-1752, 1739-1753, 1740-1754, 1741-1755, 1742-1756,
1743-1757, 1744-1758, 1745-1759, 1746-1760, 1747-1761, 1748-1762,
1749-1763, 1750-1764, 1751-1765, 1752-1766,
1753-1767, 1754-1768, 1755-1769, 1756-1770, 1757-1771, 1758-1772,
1759-1773, 1760-1774, 1761-1775, 1762-1776, 1763-1777, 1764-1778,
1765-1779, 1766-1780, 1767-1781, 1768-1782, 1769-1783, 1770-1784,
1771-1785, 1772-1786, 1773-1787, 1774-1788, 1775-1789, 1776-1790,
1777-1791, 1778-1792, 1779-1793, 1780-1794, 1781-1795, 1782-1796,
1783-1797, 1784-1798, 1785-1799, 1786-1800, 1787-1801, 1788-1802,
1789-1803, 1790-1804, 1791-1805, 1792-1806, 1793-1807, 1794-1808,
1795-1809, 1796-1810, 1797-1811, 1798-1812, 1799-1813, 1800-1814,
1801-1815, 1802-1816, 1803-1817, 1804-1818, 1805-1819, 1806-1820,
1807-1821, 1808-1822, 1809-1823, 1810-1824, 1811-1825, 1812-1826,
1813-1827, 1814-1828, 1815-1829, 1816-1830, 1817-1831, 1818-1832,
1819-1833, 1820-1834, 1821-1835, 1822-1836, 1823-1837, 1824-1838,
1825-1839, 1826-1840, 1827-1841, 1828-1842, 1829-1843, 1830-1844,
1831-1845, 1832-1846, 1833-1847, 1834-1848, 1835-1849, 1836-1850,
1837-1851, 1838-1852, 1839-1853, 1840-1854, 1841-1855, 1842-1856,
1843-1857, 1844-1858, 1845-1859, 1846-1860, 1847-1861, 1848-1862,
1849-1863, 1850-1864, 1851-1865, 1852-1866, 1853-1867, 1854-1868,
1855-1869, 1856-1870, 1857-1871, 1858-1872, 1859-1873, 1860-1874,
1861-1875, 1862-1876, 1863-1877, 1864-1878, 1865-1879, 1866-1880,
1867-1881, 1868-1882, 1869-1883, 1870-1884, 1871-1885, 1872-1886,
1873-1887, 1874-1888, 1875-1889, 1876-1890, 1877-1891, 1878-1892,
1879-1893, 1880-1894, 1881-1895, 1882-1896, 1883-1897, 1884-1898,
1885-1899, 1886-1900, 1887-1901, 1888-1902, 1889-1903, 1890-1904,
1891-1905, 1892-1906, 1893-1907, 1894-1908, 1895-1909, 1896-1910,
1897-1911, 1898-1912, 1899-1913, 1900-1914, 1901-1915, 1902-1916,
1903-1917, 1904-1918, 1905-1919, 1906-1920, 1907-1921, 1908-1922,
1909-1923, 1910-1924, 1911-1925, 1912-1926, 1913-1927, 1914-1928,
1915-1929, 1916-1930, 1917-1931, 1918-1932, 1919-1933, 1920-1934,
1921-1935, 1922-1936, 1923-1937, 1924-1938, 1925-1939, 1926-1940,
1927-1941, 1928-1942, 1929-1943, 1930-1944, 1931-1945, 1932-1946,
1933-1947, 1934-1948, 1935-1949, 1936-1950, 1937-1951, 1938-1952,
1939-1953, 1940-1954, 1941-1955, 1942-1956, 1943-1957, 1944-1958,
1945-1959, 1946-1960, 1947-1961, 1948-1962, 1949-1963, 1950-1964,
1951-1965, 1952-1966, 1953-1967, 1954-1968, 1955-1969, 1956-1970,
1957-1971, 1958-1972, 1959-1973, 1960-1974, 1961-1975, 1962-1976,
1963-1977, 1964-1978, 1965-1979, 1966-1980, 1967-1981, 1968-1982,
1969-1983, 1970-1984, 1971-1985, 1972-1986, 1973-1987, 1974-1988,
1975-1989, 1976-1990, 1977-1991, 1978-1992, 1979-1993, 1980-1994,
1981-1995, 1982-1996, 1983-1997, 1984-1998, 1985-1999, 1986-2000,
1987-2001, 1988-2002, 1989-2003, 1990-2004, 1991-2005, 1992-2006,
1993-2007, 1994-2008, 1995-2009, 1996-2010, 1997-2011, 1998-2012,
1999-2013, 2000-2014, 2001-2015, 2002-2016, 2003-2017, 2004-2018,
2005-2019, 2006-2020, 2007-2021, 2008-2022, 2009-2023, 2010-2024,
2011-2025, 2012-2026, 2013-2027, 2014-2028, 2015-2029, 2016-2030,
2017-2031, 2018-2032, 2019-2033, 2020-2034, 2021-2035, 2022-2036,
2023-2037, 2024-2038, 2025-2039, 2026-2040, 2027-2041, 2028-2042,
2029-2043, 2030-2044, 2031-2045, 2032-2046, 2033-2047, 2034-2048,
2035-2049, 2036-2050, 2037-2051, 2038-2052, 2039-2053, 2040-2054,
2041-2055, 2042-2056, 2043-2057, 2044-2058, 2045-2059, 2046-2060,
2047-2061, 2048-2062, 2049-2063, 2050-2064, 2051-2065, 2052-2066,
2053-2067, 2054-2068, 2055-2069, 2056-2070, 2057-2071, 2058-2072,
2059-2073, 2060-2074, 2061-2075, 2062-2076, 2063-2077, 2064-2078,
2065-2079, 2066-2080, 2067-2081, 2068-2082, 2069-2083, 2070-2084,
2071-2085, 2072-2086, 2073-2087, 2074-2088, 2075-2089, 2076-2090,
2077-2091, 2078-2092, 2079-2093, 2080-2094, 2081-2095, 2082-2096,
2083-2097, 2084-2098, 2085-2099, 2086-2100, 2087-2101, 2088-2102,
2089-2103, 2090-2104, 2091-2105, 2092-2106, 2093-2107, 2094-2108,
2095-2109, 2096-2110, 2097-2111, 2098-2112, 2099-2113, 2100-2114,
2101-2115, 2102-2116, 2103-2117, 2104-2118, 2105-2119, 2106-2120,
2107-2121, 2108-2122, 2109-2123, 2110-2124, 2111-2125, 2112-2126,
2113-2127, 2114-2128, 2115-2129, 2116-2130, 2117-2131, 2118-2132,
2119-2133, 2120-2134, 2121-2135, 2122-2136, 2123-2137, 2124-2138,
2125-2139, 2126-2140, 2127-2141, 2128-2142, 2129-2143, 2130-2144,
2131-2145, 2132-2146, 2133-2147, 2134-2148, 2135-2149, 2136-2150,
2137-2151, 2138-2152, 2139-2153, 2140-2154, 2141-2155, 2142-2156,
2143-2157, 2144-2158, 2145-2159, 2146-2160, 2147-2161, 2148-2162,
2149-2163, 2150-2164, 2151-2165, 2152-2166, 2153-2167, 2154-2168,
2155-2169, 2156-2170, 2157-2171, 2158-2172, 2159-2173, 2160-2174,
2161-2175, 2162-2176, 2163-2177, 2164-2178, 2165-2179, 2166-2180,
2167-2181, 2168-2182, 2169-2183, 2170-2184, 2171-2185, 2172-2186,
2173-2187, 2174-2188, 2175-2189, 2176-2190, 2177-2191, 2178-2192,
2179-2193, 2180-2194, 2181-2195, 2182-2196, 2183-2197, 2184-2198,
2185-2199, 2186-2200, 2187-2201, 2188-2202, 2189-2203, 2190-2204,
2191-2205, 2192-2206, 2193-2207, 2194-2208, 2195-2209, 2196-2210,
2197-2211, 2198-2212, 2199-2213, 2200-2214, 2201-2215, 2202-2216,
2203-2217, 2204-2218, 2205-2219, 2206-2220, 2207-2221, 2208-2222,
2209-2223, 2210-2224, 2211-2225, 2212-2226, 2213-2227, 2214-2228,
2215-2229, 2216-2230, 2217-2231, 2218-2232, 2219-2233, 2220-2234,
2221-2235, 2222-2236, 2223-2237, 2224-2238, 2225-2239, 2226-2240,
2227-2241, 2228-2242, 2229-2243, 2230-2244, 2231-2245, 2232-2246,
2233-2247, 2234-2248, 2235-2249, 2236-2250, 2237-2251, 2238-2252,
2239-2253, 2240-2254, 2241-2255, 2242-2256, 2243-2257, 2244-2258,
2245-2259, 2246-2260, 2247-2261, 2248-2262, 2249-2263, 2250-2264,
2251-2265, 2252-2266, 2253-2267, 2254-2268, 2255-2269, 2256-2270,
2257-2271, 2258-2272, 2259-2273, 2260-2274, 2261-2275, 2262-2276,
2263-2277, 2264-2278, 2265-2279, 2266-2280, 2267-2281, 2268-2282,
2269-2283, 2270-2284, 2271-2285, 2272-2286, 2273-2287, 2274-2288,
2275-2289, 2276-2290, 2277-2291, 2278-2292, 2279-2293, 2280-2294,
2281-2295, 2282-2296, 2283-2297, 2284-2298, 2285-2299, 2286-2300,
2287-2301, 2288-2302, 2289-2303, 2290-2304, 2291-2305, 2292-2306,
2293-2307, 2294-2308, 2295-2309, 2296-2310, 2297-2311, 2298-2312,
2299-2313, 2300-2314, 2301-2315, 2302-2315, 2303-2317, 2304-2318,
2305-2319, 2306-2320, 2307-2321, 2308-2322, 2309-2323, 2310-2324,
2311-2325, 2312-2326, 2313-2327, 2314-2328, 2315-2329, 2316-2330,
2317-2331, 2318-2332, 2319-2333, 2320-2334, 2321-2335, 2322-2336,
2323-2337, 2324-2338, 2325-2339, 2326-2340, 2327-2341, 2328-2342,
2329-2343, 2330-2344, 2331-2345, 2332-2346, 2333-2347, 2334-2348,
2335-2349, 2336-2350, 2337-2351, 2338-2352, 2339-2353, 2340-2354,
2341-2355, 2342-2356, 2343-2357, 2344-2358, 2345-2359, 2346-2360,
2347-2361, 2348-2362, 2349-2363, 2350-2364, 2351-2365, 2352-2366,
2353-2367, 2354-2368, 2355-2369, 2356-2370, 2357-2371, 2358-2372,
2359-2373, 2360-2374, 2361-2375, 2362-2376, 2363-2377, 2364-2378,
2365-2379, 2366-2380, 2367-2381, 2368-2382, 2369-2383, 2370-2384,
2371-2385, 2372-2386, 2373-2387, 2374-2388, 2375-2389, 2376-2390,
2377-2391, 2378-2392, 2379-2393, 2380-2394, 2381-2395, 2382-2396,
2383-2397, 2384-2398, 2385-2399, 2386-2400, 2387-2401, 2388-2402,
2389-2403, 2390-2404, 2391-2405, 2392-2406, 2393-2407, 2394-2408,
2395-2409, 2396-2410, 2397-2411, 2398-2412, 2399-2413, 2400-1412,
2401-2415, 2402-2416, 2403-2417, 2404-2418, 2405-2419, 2406-2420,
2407-2421, 2408-2422, 2409-2423, 2410-2424, 2411-2425, 2412-2426,
2413-2427, 2414-2428, 2415-2429, 2416-2430, 2417-2431, 2418-2432,
2419-2433, 2420-2434, 2421-2435, 2422-2436, 2423-2437, 2424-2438,
2425-2439, 2426-2440, 2427-2441, 2428-2442, 2429-2443, 2430-2444,
2431-2445, 2432-2446, 2433-2447, 2434-2448, 2435-2449, 2436-2450,
2437-2451, 2438-2452, 2439-2453, 2440-2454, 2441-2455, 2442-2456,
2443-2457, 2444-2458, 2445-2459, 2446-2460, 2447-2461, 2448-2462,
2449-2463, 2450-2464, 2451-2465, 2452-2466, 2453-2467, 2454-2468,
2455-2469, 2456-2470, 2457-2471, 2458-2472, 2459-2473, 2460-2474,
2461-2475, 2462-2476, 2463-2477, 2464-2478, 2465-2479, 2466-2480,
2467-2481, 2468-2482, 2469-2483, 2470-2484, 2471-2485, 2472-2486,
2473-2487, 2474-2488, 2475-2489, 2476-2490, 2477-2491, 2478-2492,
2479-2493, 2480-2494, 2481-2495, 2482-2496, 2483-2497, 2484-2498,
2485-2499, 2486-2500, 2487-2501, 2488-2502, 2489-2503, 2490-2504,
2491-2505, 2492-2506, 2493-2507, 2494-2508, 2495-2509, 2496-2510,
2497-2511, 2498-2513, 2499-2514, 2500-2515, 2501-2515, 2502-2516,
2503-2517, 2504-2518, 2505-2519, 2506-2520, 2507-2521, 2508-2522,
2509-2523, 2510-2524, 2511-2525, 2512-2526, 2513-2527, 2514-2528,
2515-2529, 2516-2530, 2517-2531, 2518-2532, 2519-2533, 2520-2534,
2521-2535, 2522-2536, 2523-2537, 2524-2538, 2525-2539, 2526-2540,
2527-2541, 2528-2542, 2529-2543, 2530-2544, 2531-2545, 2532-2546,
2533-2547, 2534-2548, 2535-2549, 2536-2550, 2537-2551, 2538-2552,
2539-2553, 2540-2554, 2541-2555, 2542-2556, 2543-2557, 2544-2558,
2545-2559, 2546-2560, 2547-2561, 2548-2562, 2549-2563, 2550-2564,
2551-2565, 2552-2566, 2553-2567, 2554-2568, 2555-2569, 2556-2570,
2557-2571, 2558-2572, 2559-2573, 2560-2574, 2561-2575, 2562-2576,
2563-2577, 2564-2578, 2565-2579, 2566-2580, 2567-2581, 2568-2582,
2569-2583, 2570-2584, 2571-2585, 2572-2586, 2573-2587, 2574-2588,
2575-2589, 2576-2590, 2577-2591, 2578-2592, 2579-2593, 2580-2594,
2581-2595, 2582-2596, 2583-2597, 2584-2598, 2585-2599, 2586-2600,
2587-2601, 2588-2602, 2589-2603, 2590-2604, 2591-2605, 2592-2606,
2593-2607, 2594-2608, 2595-2609, 2596-2610, 2597-2611, 2598-2612,
2599-2613, 2600-2614, 2601-2615, 2602-2616, 2603-2617, 2604-2618,
2605-2619, 2606-2620, 2607-2621, 2608-2622, 2609-2623, 2610-2624,
2611-2625, 2612-2626, 2613-2627, 2614-2628, 2615-2629, 2616-2630,
2617-2631, 2618-2632, 2619-2633, 2620-2634, 2621-2635, 2622-2636,
2623-2637, 2624-2638, 2625-2639, 2626-2640, 2627-2641, 2628-2642,
2629-2643, 2630-2644, 2631-2645, 2632-2646, 2633-2647, 2634-2648,
2635-2649, 2636-2650, 2637-2651, 2638-2652, 2639-2653, 2640-2654,
2641-2655, 2642-2656, 2643-2657, 2644-2658, 2645-2659, 2646-2660,
2647-2661, 2648-2662, 2649-2663, 2650-2664, 2651-2665, 2652-2666,
2653-2667, 2654-2668, 2655-2669, 2656-2670, 2657-2671, 2658-2672,
2659-2673, 2660-2674, 2661-2675, 2662-2676, 2663-2677, 2664-2678,
2665-2679, 2666-2680, 2667-2681, 2668-2682, 2669-2683, 2670-2684,
2671-2685, 2672-2686, 2673-2687, 2674-2688, 2675-2689, 2676-2690,
2677-2691, 2678-2692, 2679-2693, 2680-2694, 2681-2695, 2682-2696,
2683-2697, 2684-2698, 2685-2699, 2686-2700, 2687-2701, 2688-2702,
2689-2703, 2690-2704, 2691-2705, 2692-2706, 2693-2707, 2694-2708,
2695-2709, 2696-2710, 2697-2711, 2698-2712, 2699-2713, 2700-2714,
2701-2715, 2702-2716, 2703-2717, 2704-2718, 2705-2719, 2706-2720,
2707-2721, 2708-2722, 2709-2723, 2710-2724, 2711-2725, 2712-2726,
2713-2727, 2714-2728, 2715-2729, 2716-2730, 2717-2731, 2718-2732,
2719-2733, 2720-2734, 2721-2735, 2722-2736, 2723-2737, 2724-2738,
2725-2739, 2726-2740, 2727-2741, 2728-2742, 2729-2743, 2730-2744,
2731-2745, 2732-2746, 2733-2747, 2734-2748, 2735-2749, 2736-2750,
2737-2751, 2738-2752, 2739-2753, 2740-2754, 2741-2755, 2742-2756,
2743-2757, 2744-2758, 2745-2759, 2746-2760, 2747-2761, 2748-2762,
2749-2763, 2750-2764, 2751-2765, 2752-2766, 2753-2767, 2754-2768,
2755-2769, 2756-2770, 2757-2771, 2758-2772, 2759-2773, 2760-2774,
2761-2775, 2762-2776, 2763-2777, 2764-2778, 2765-2779, 2766-2780,
2767-2781, 2768-2782, 2769-2783, 2770-2784, 2771-2785, 2772-2786,
2773-2787, 2774-2788, 2775-2789, 2776-2790, 2777-2791, 2778-2792,
2779-2793, 2780-2794, 2781-2795, 2782-2796, 2783-2797, 2784-2798,
2785-2799, 2786-2800, 2787-2801, 2788-2802, 2789-2803, 2790-2804,
2791-2805, 2792-2806, 2793-2807, 2794-2808, 2795-2809, 2796-2810,
2797-2811, 2798-2812, 2799-2813, 2800-2814, 2801-2815, 2802-2816,
2803-2817, 2804-2818, 2805-2819, 2806-2820, 2807-2821, 2808-2822,
2809-2823, 2810-2824, 2811-2825, 2812-2826, 2813-2827, 2814-2828,
2815-2829, 2816-2830, 2817-2831, 2818-2832, 2819-2833, 2820-2834,
2821-2835, 2822-2836, 2823-2837, 2824-2838, 2825-2839, 2826-2840,
2827-2841, 2828-2842, 2829-2843, 2830-2844, 2831-2845, 2832-2846,
2833-2847, 2834-2848, 2835-2849, 2836-2850, 2837-2851, 2838-2852,
2839-2853, 2840-2854, 2841-2855, 2842-2856, 2843-2857, 2844-2858,
2845-2859, 2846-2860, 2847-2861, 2848-2862, 2849-2863, 2850-2864,
2851-2865, 2852-2866, 2853-2867, 2854-2868, 2855-2869, 2856-2870,
2857-2871, 2858-2872, 2859-2873, 2860-2874, 2861-2875, 2862-2876,
2863-2877, 2864-2878, 2865-2879, 2866-2880, 2867-2881, 2868-2882,
2869-2883, 2870-2884, 2871-2885, 2872-2886, 2873-2887, 2874-2888,
2875-2889, 2876-2890, 2877-2891, 2878-2892, 2879-2893, 2880-2894,
2881-2895, 2882-2896, 2883-2897, 2884-2898, 2885-2899, 2886-2900,
2887-2901, 2888-2902, 2889-2903, 2890-2904, 2891-2905, 2892-2906,
2893-2907, 2894-2908, 2895-2909, 2896-2910, 2897-2911, 2898-2912,
2899-2913, 2900-2914, 2901-2915, 2902-2916, 2903-2917, 2904-2918,
2905-2919, 2906-2920, 2907-2921, 2908-2922, 2909-2923, 2910-2924,
2911-2925, 2912-2926, 2913-2927, 2914-2928, 2915-2929, 2916-2930,
2917-2931, 2918-2932, 2919-2933, 2920-2934, 2921-2935, 2922-2936,
2923-2937, 2924-2938, 2925-2939, 2926-2940, 2927-2941, 2928-2942,
2929-2943, 2930-2944, 2931-2945, 2932-2946, 2933-2947, 2934-2948,
2935-2949, 2936-2950, 2937-2951, 2938-2952, 2939-2953, 2940-2954,
2941-2955, 2942-2956, 2943-2957, 2944-2958, 2945-2959, 2946-2960,
2947-2961, 2948-2962, 2949-2963, 2950-2964, 2951-2965, 2952-2966,
2953-2967, 2954-2968, 2955-2969, 2956-2970, 2957-2971, 2958-2972,
2959-2973, 2960-2974, 2961-2975, 2962-2976, 2963-2977, 2964-2978,
2965-2979, 2966-2980, 2967-2981, 2968-2982, 2969-2983, 2970-2984,
2971-2985, 2972-2986, 2973-2987, 2974-2988, 2975-2989, 2976-2990,
2977-2991, 2978-2992, 2979-2993, 2980-2994, 2981-2995, 2982-2996,
2983-2997, 2984-2998, 2985-2999, 2986-3000, 2987-3001, 2988-3002,
2989-3003, 2990-3004, 2991-3005, 2992-3006, 2993-3007, 2994-3008,
2995-3009, 2996-3010, 2997-3011, 2998-3012, 2999-3013, 3000-3014,
3001-3015, 3002-3016, 3003-3017, 3004-3018, 3005-3019, 3006-3020,
3007-3021, 3008-3022, 3009-3023, 3010-3024, 3011-3025, 3012-3026,
3013-3027, 3014-3028, 3015-3029, 3016-3030, 3017-3031, 3018-3032,
3019-3033, 3020-3034, 3021-3035, 3022-3036, 3023-3037, 3024-3038,
3025-3039, 3026-3040, 3027-3041, 3028-3042, 3029-3043, 3030-3044,
3031-3045, 3032-3046, 3033-3047, 3034-3048, 3035-3049, 3036-3050,
3037-3051, 3038-3052, 3039-3053, 3040-3054, 3041-3055, 3042-3056,
3043-3057, 3044-3058, 3045-3059, 3046-3060, 3047-3061, 3048-3062,
3049-3063, 3050-3064, 3051-3065, 3052-3066, 3053-3067, 3054-3068,
3055-3069, 3056-3070, 3057-3071, 3058-3072, 3059-3073, 3060-3074,
3061-3075, 3062-3076, 3063-3077, 3064-3078, 3065-3079, 3066-3080,
3067-3081, 3068-3082, 3069-3083, 3070-3084, 3071-3085, 3072-3086,
3073-3087, 3074-3088, 3075-3089, 3076-3090, 3077-3091, 3078-3092,
3079-3093, 3080-3094, 3081-3095, 3082-3096, 3083-3097, 3084-3098,
3085-3099, 3086-3100, 3087-3101, 3088-3102, 3089-3103, 3090-3104,
3091-3105, 3092-3106, 3093-3107, 3094-3108, 3095-3109, 3096-3110,
3097-3111, 3098-3112, 3099-3113, 3100-3114, 3101-3115, 3102-3116,
3103-3117, 3104-3118, 3105-3119, 3106-3120, 3107-3121, 3108-3122,
3109-3123, 3110-3124, 3111-3125, 3112-3126, 3113-3127, 3114-3128,
3115-3129, 3116-3130, 3117-3131, 3118-3132, 3119-3133, 3120-3134,
3121-3135, 3122-3136, 3123-3137, 3124-3138, 3125-3139, 3126-3140,
3127-3141, 3128-3142, 3129-3143, 3130-3144, 3131-3145, 3132-3146,
3133-3147, 3134-3148, 3135-3149, 3136-3150, 3137-3151, 3138-3152,
3139-3153, 3140-3154, 3141-3155, 3142-3156, 3143-3157, 3144-3158,
3145-3159, 3146-3160, 3147-3161, 3148-3162, 3149-3163, 3150-3164,
3151-3165, 3152-3166, 3153-3167, 3154-3168, 3155-3169, 3156-3170,
3157-3171, 3158-3172, 3159-3173, 3160-3174, 3161-3175, 3162-3176,
3163-3177, 3164-3178, 3165-3179, 3166-3180, 3167-3181, 3168-3182,
3169-3183, 3170-3184, 3171-3185, 3172-3186, 3173-3187, 3174-3188,
3175-3189, 3176-3190, 3177-3191, 3178-3192, 3179-3193, 3180-3194,
3181-3195, 3182-3196, 3183-3197, 3184-3198, 3185-3199, 3186-3200,
3187-3201, 3188-3202, 3189-3203, 3190-3204, 3191-3205, 3192-3206,
3193-3207, 3194-3208, 3195-3209, 3196-3210, 3197-3211, 3198-3212,
3199-3213, 3200-3214, 3201-3215, 3202-3216, 3203-3217, 3204-3218,
3205-3219, 3206-3220, 3207-3221, 3208-3222, 3209-3223, 3210-3224,
3211-3225, 3212-3226, 3213-3227, 3214-3228, 3215-3229, 3216-3230,
3217-3231, 3218-3232, 3219-3233, 3220-3234, 3221-3235, 3222-3236,
3223-3237, 3224-3238, 3225-3239, 3226-3240, 3227-3241, 3228-3242,
3229-3243, 3230-3244, 3231-3245, 3232-3246, 3233-3247, 3234-3248,
3235-3249, 3236-3250, 3237-3251, 3238-3252, 3239-3253, 3240-3254,
3241-3255, 3242-3256, 3243-3257, 3244-3258, 3245-3259, 3246-3260,
3247-3261, 3248-3262, 3249-3263, 3250-3264, 3251-3265, 3252-3266,
3253-3267, 3254-3268, 3255-3269, 3256-3270, 3257-3271,
3258-3272,
3259-3273, 3260-3274, 3261-3275, 3262-3276, 3263-3277, 3264-3278,
3265-3279, 3266-3280, 3267-3281, 3268-3282, 3269-3283, 3270-3284,
3271-3285, 3272-3286, 3273-3287, 3274-3288, 3275-3289, 3276-3290,
3277-3291, 3278-3292, 3279-3293, 3280-3294, 3281-3295, 3282-3296,
3283-3297, 3284-3298, 3285-3299, 3286-3300, 3287-3301, 3288-3302,
3289-3303, 3290-3304, 3291-3305, 3292-3306, 3293-3307, 3294-3308,
3295-3309, 3296-3310, 3297-3311, 3298-3312, 3299-3313, 3300-3314,
3301-3315, 3302-3315, 3303-3317, 3304-3318, 3305-3319, 3306-3320,
3307-3321, 3308-3322, 3309-3323, 3310-3324, 3311-3325, 3312-3326,
3313-3327, 3314-3328, 3315-3329, 3316-3330, 3317-3331, 3318-3332,
3319-3333, 3320-3334, 3321-3335, 3322-3336, 3323-3337, 3324-3338,
3325-3339, 3326-3340, 3327-3341, 3328-3342, 3329-3343, 3330-3344,
3331-3345, 3332-3346, 3333-3347, 3334-3348, 3335-3349, 3336-3350,
3337-3351, 3338-3352, 3339-3353, 3340-3354, 3341-3355, 3342-3356,
3343-3357, 3344-3358, 3345-3359, 3346-3360, 3347-3361, 3348-3362,
3349-3363, 3350-3364, 3351-3365, 3352-3366, 3353-3367, 3354-3368,
3355-3369, 3356-3370, 3357-3371, 3358-3372, 3359-3373, 3360-3374,
3361-3375, 3362-3376, 3363-3377, 3364-3378, 3365-3379, 3366-3380,
3367-3381, 3368-3382, 3369-3383, 3370-3384, 3371-3385, 3372-3386,
3373-3387, 3374-3388, 3375-3389, 3376-3390, 3377-3391, 3378-3392,
3379-3393, 3380-3394, 3381-3395, 3382-3396, 3383-3397, 3384-3398,
3385-3399, 3386-3400, 3387-3401, 3388-3402, 3389-3403, 3390-3404,
3391-3405, 3392-3406, 3393-3407, 3394-3408, 3395-3409, 3396-3410,
3397-3411, 3398-3412, 3399-3413, 3400-3414, 3401-3415, 3402-3416,
3403-3417, 3404-3418, 3405-3419, 3406-3420, 3407-3421, 3408-3422,
3409-3423, 3410-3424, 3411-3425, 3412-3426, 3413-3427, 3414-3428,
3415-3429, 3416-3430, 3417-3431, 3418-3432, 3419-3433, 3420-3434,
3421-3435, 3422-3436, 3423-3437, 3424-3438, 3425-3439, 3426-3440,
3427-3441, 3428-3442, 3429-3443, 3430-3444, 3431-3445, 3432-3446,
3433-3447, 3434-3448, 3435-3449, 3436-3450, 3437-3451, 3438-3452,
3439-3453, 3440-3454, 3441-3455, 3442-3456, 3443-3457, 3444-3458,
3445-3459, 3446-3460, 3447-3461, 3448-3462, 3449-3463, 3450-3464,
3451-3465, 3452-3466, 3453-3467, 3454-3468, 3455-3469, 3456-3470,
3457-3471, 3458-3472, 3459-3473, 3460-3474, 3461-3475, 3462-3476,
3463-3477, 3464-3478, 3465-3479, 3466-3480, 3467-3481, 3468-3482,
3469-3483, 3470-3484, 3471-3485, 3472-3486, 3473-3487, 3474-3488,
3475-3489, 3476-3490, 3477-3491, 3478-3492, 3479-3493, 3480-3494,
3481-3495, 3482-3496, 3483-3497, 3484-3498, 3485-3499, 3486-3500,
3487-3501, 3488-3502, 3489-3503, 3490-3504, 3491-3505, 3492-3506,
3493-3507, 3494-3508, 3495-3509, 3496-3510, 3497-3511, 3498-3513,
3499-3514, 3500-3515, 3501-3515, 3502-3516, 3503-3517, 3504-3518,
3505-3519, 3506-3520, 3507-3521, 3508-3522, 3509-3523, 3510-3524,
3511-3525, 3512-3526, 3513-3527, 3514-3528, 3515-3529,
3516-3530,
3517-3531, 3518-3532, 3519-3533, 3520-3534, 3521-3535, 3522-3536,
3523-3537, 3524-3538, 3525-3539, 3526-3540, 3527-3541, 3528-3542,
3529-3543, 3530-3544, 3531-3545, 3532-3546, 3533-3547, 3534-3548,
3535-3549, 3536-3550, 3537-3551, 3538-3552, 3539-3553, 3540-3554,
3541-3555, 3542-3556, 3543-3557, 3544-3558, 3545-3559, 3546-3560,
3547-3561, 3548-3562, 3549-3563, 3550-3564, 3551-3565, 3552-3566,
3553-3567, 3554-3568, 3555-3569, 3556-3570, 3557-3571, 3558-3572,
3559-3573, 3560-3574, 3561-3575, 3562-3576, 3563-3577, 3564-3578,
3565-3579, 3566-3580, 3567-3581, 3568-3582, 3569-3583, 3570-3584,
3571-3585, 3572-3586, 3573-3587, 3574-3588, 3575-3589, 3576-3590,
3577-3591, 3578-3592, 3579-3593, 3580-3594, 3581-3595, 3582-3596,
3583-3597, 3584-3598, 3585-3599, 3586-3600, 3587-3601, 3588-3602,
3589-3603, 3590-3604, 3591-3605, 3592-3606, 3593-3607, 3594-3608,
3595-3609, 3596-3610, 3597-3611, 3598-3612, 3599-3613, 3600-3614,
3601-3615, 3602-3616, 3603-3617, 3604-3618, 3605-3619, 3606-3620,
3607-3621, 3608-3622, 3609-3623, 3610-3624, 3611-3625, 3612-3626,
3613-3627, 3614-3628, 3615-3629, 3616-3630, 3617-3631, 3618-3632,
3619-3633, 3620-3634, 3621-3635, 3622-3636, 3623-3637, 3624-3638,
3625-3639, 3626-3640, 3627-3641, 3628-3642, 3629-3643, 3630-3644,
3631-3645, 3632-3646, 3633-3647, 3634-3648, 3635-3649, 3636-3650,
3637-3651, 3638-3652, 3639-3653, 3640-3654, 3641-3655, 3642-3656,
3643-3657, 3644-3658, 3645-3659, 3646-3660, 3647-3661, 3648-3662,
3649-3663, 3650-3664, 3651-3665, 3652-3666, 3653-3667, 3654-3668,
3655-3669, 3656-3670, 3657-3671, 3658-3672, 3659-3673, 3660-3674,
3661-3675, 3662-3676, 3663-3677, 3664-3678, 3665-3679, 3666-3680,
3667-3681, 3668-3682, 3669-3683, 3670-3684, 3671-3685, 3672-3686,
3673-3687, 3674-3688, 3675-3689, 3676-3690, 3677-3691, 3678-3692,
3679-3693, 3680-3694, 3681-3695, 3682-3696, 3683-3697, 3684-3698,
3685-3699, 3686-3700, 3687-3701, 3688-3702, 3689-3703, 3690-3704,
3691-3705, 3692-3706, 3693-3707, 3694-3708, 3695-3709, 3696-3710,
3697-3711, 3698-3712, 3699-3713, 3700-3701, 3701-3715, 3702-3716,
3703-3717, 3704-3718, 3705-3719, 3706-3720, 3707-3721, 3708-3722,
3709-3723, 3710-3724, 3711-3725, 3712-3726, 3713-3727, 3714-3728,
3715-3729, 3716-3730, 3717-3731, 3718-3732, 3719-3733, 3720-3734,
3721-3735, 3722-3736, 3723-3737, 3724-3738, 3725-3739, 3726-3740,
3727-3741, 3728-3742, 3729-3743, 3730-3744, 3731-3745, 3732-3746,
3733-3747, 3734-3748, 3735-3749, 3736-3750, 3737-3751, 3738-3752,
3739-3753, 3740-3754, 3741-3755, 3742-3756, 3743-3757, 3744-3758,
3745-3759, 3746-3760, 3747-3761, 3748-3762, 3749-3763, 3750-3764,
3751-3765, 3752-3766, 3753-3767, 3754-3768, 3755-3769, 3756-3770,
3757-3771, 3758-3772, 3759-3773, 3760-3774, 3761-3775, 3762-3776,
3763-3777, 3764-3778, 3765-3779, 3766-3780, 3767-3781, 3768-3782,
3769-3783, 3770-3784, 3771-3785, 3772-3786, 3773-3787, 3774-3788,
3775-3789, 3776-3790, 3777-3791, 3778-3792, 3779-3793, 3780-3794,
3781-3795, 3782-3796, 3783-3797, 3784-3798, 3785-3799, 3786-3800,
3787-3801, 3788-3802, 3789-3803, 3790-3804, 3791-3805, 3792-3806,
3793-3807, 3794-3808, 3795-3809, 3796-3810, 3797-3811, 3798-3812,
3799-3813, 3800-3814, 3801-3815, 3802-3816, 3803-3817, 3804-3818,
3805-3819, 3806-3820, 3807-3821, 3808-3822, 3809-3823, 3810-3824,
3811-3825, 3812-3826, 3813-3827, 3814-3828, 3815-3829, 3816-3830,
3817-3831, 3818-3832, 3819-3833, 3820-3834, 3821-3835, 3822-3836,
3823-3837, 3824-3838, 3825-3839, 3826-3840, 3827-3841, 3828-3842,
3829-3843, 3830-3844, 3831-3845, 3832-3846, 3833-3847, 3834-3848,
3835-3849, 3836-3850, 3837-3851, 3838-3852, 3839-3853, 3840-3854,
3841-3855, 3842-3856, 3843-3857, 3844-3858, 3845-3859, 3846-3860,
3847-3861, 3848-3862, 3849-3863, 3850-3864, 3851-3865, 3852-3866,
3853-3867, 3854-3868, 3855-3869, 3856-3870, 3857-3871, 3858-3872,
3859-3873, 3860-3874, 3861-3875, 3862-3876, 3863-3877, 3864-3878,
3865-3879, 3866-3880, 3867-3881, 3868-3882, 3869-3883, 3870-3884,
3871-3885, 3872-3886, 3873-3887, 3874-3888, 3875-3889, 3876-3890,
3877-3891, 3878-3892, 3879-3893, 3880-3894, 3881-3895, 3882-3896,
3883-3897, 3884-3898, 3885-3899, 3886-3900, 3887-3901, 3888-3902,
3889-3903, 3890-3904, 3891-3905, 3892-3906, 3893-3907, 3894-3908,
3895-3909, 3896-3910, 3897-3911, 3898-3912, 3899-3913, 3900-3914,
3901-3915, 3902-3916, 3903-3917, 3904-3918, 3905-3919, 3906-3920,
3907-3921, 3908-3922, 3909-3923, 3910-3924, 3911-3925, 3912-3926,
3913-3927, 3914-3928, 3915-3929, 3916-3930, 3917-3931, 3918-3932,
3919-3933, 3920-3934, 3921-3935, 3922-3936, 3923-3937, 3924-3938,
3925-3939, 3926-3940, 3927-3941, 3928-3942, 3929-3943, 3930-3944,
3931-3945, 3932-3946, 3933-3947, 3934-3948, 3935-3949, 3936-3950,
3937-3951, 3938-3952, 3939-3953, 3940-3954, 3941-3955, 3942-3956,
3943-3957, 3944-3958, 3945-3959, 3946-3960, 3947-3961, 3948-3962,
3949-3963, 3950-3964, 3951-3965, 3952-3966, 3953-3967, 3954-3968,
3955-3969, 3956-3970, 3957-3971, 3958-3972, 3959-3973, 3960-3974,
3961-3975, 3962-3976, 3963-3977, 3964-3978, 3965-3979, 3966-3980,
3967-3981, 3968-3982, 3969-3983, 3970-3984, 3971-3985, 3972-3986,
3973-3987, 3974-3988, 3975-3989, 3976-3990, 3977-3991, 3978-3992,
3979-3993, 3980-3994, 3981-3995, 3982-3996, 3983-3997, 3984-3998,
3985-3999, 3986-4000, 3987-4001, 3988-4002, 3989-4003, 3990-4004,
3991-4005, 3992-4006, 3993-4007, 3994-4008, 3995-4009, 3996-4010,
3997-4011, 3998-4012, 3999-4013, 4000-4914, 4001-4015, 4002-4016,
4003-4017, 4004-4018, 4005-4019, 4006-4020, 4007-4021, 4008-4022,
4009-4023, 4010-4024, 4011-4025, 4012-4026, 4013-4027, 4014-4028,
4015-4029, 4016-4030, 4017-4031, 4018-4032, 4019-4033, 4020-4034,
4021-4035, 4022-4036, 4023-4037, 4024-4038, 4025-4039, 4026-4040,
4027-4041, 4028-4042, 4029-4043, 4030-4044, 4031-4045, 4032-4046,
4033-4047, 4034-4048, 4035-4049, 4036-4050, 4037-4051, 4038-4052,
4039-4053, 4040-4054, 4041-4055, 4042-4056, 4043-4057, 4044-4058,
4045-4059, 4046-4060, 4047-4061, 4048-4062, 4049-4063, 4050-4064,
4051-4065, 4052-4066, 4053-4067, 4054-4068, 4055-4069, 4056-4070,
4057-4071, 4058-4072, 4059-4073, 4060-4074, 4061-4075, 4062-4076,
4063-4077, 4064-4078, 4065-4079, 4066-4080, 4067-4081, 4068-4082,
4069-4083, 4070-4084, 4071-4085, 4072-4086, 4073-4087, 4074-4088,
4075-4089, 4076-4090, 4077-4091, 4078-4092, 4079-4093, 4080-4094,
4081-4095, 4082-4096, 4083-4097, 4084-4098, 4085-4099, 4086-4100,
4087-4101, 4088-4102, 4089-4103, 4090-4104, 4091-4105, 4092-4106,
4093-4107, 4094-4108, 4095-4109, 4096-4110, 4097-4111, 4098-4112,
4099-4113, 4100-4114, 4101-4115, 4102-4116, 4103-4117, 4104-4118,
4105-4119, 4106-4120, 4107-4121, 4108-4122, 4109-4123, 4110-4124,
4111-4125, 4112-4126, 4113-4127, 4114-4128, 4115-4129, 4116-4130,
4117-4131, 4118-4132, 4119-4133, 4120-4134, 4121-4135, 4122-4136,
4123-4137, 4124-4138, 4125-4139, 4126-4140, 4127-4141, 4128-4142,
4129-4143, 4130-4144, 4131-4145, 4132-4146, 4133-4147, 4134-4148,
4135-4149, 4136-4150, 4137-4151, 4138-4152, 4139-4153, 4140-4154,
4141-4155, 4142-4156, 4143-4157, 4144-4158, 4145-4159, 4146-4160,
4147-4161, 4148-4162, 4149-4163, 4150-4164, 4151-4165, 4152-4166,
4153-4167, 4154-4168, 4155-4169, 4156-4170, 4157-4171, 4158-4172,
4159-4173, 4160-4174, 4161-4175, 4162-4176, 4163-4177, 4164-4178,
4165-4179, 4166-4180, 4167-4181, 4168-4182, 4169-4183, 4170-4184,
4171-4185, 4172-4186, 4173-4187, 4174-4188, 4175-4189, 4176-4190,
4177-4191, 4178-4192, 4179-4193, 4180-4194, 4181-4195, 4182-4196,
4183-4197, 4184-4198, 4185-4199, 4186-4200, 4187-4201, 4188-4202,
4189-4203, 4190-4204, 4191-4205, 4192-4206, 4193-4207, 4194-4208,
4195-4209, 4196-4210, 4197-4211, 4198-4212, 4199-4213, 4200-4214,
4201-4215, 4202-4216, 4203-4217, 4204-4218, 4205-4219, 4206-4220,
4207-4221, 4208-4222, 4209-4223, 4210-4224, 4211-4225, 4212-4226,
4213-4227, 4214-4228, 4215-4229, 4216-4230, 4217-4231, 4218-4232,
4219-4233, 4220-4234, 4221-4235, 4222-4236, 4223-4237, 4224-4238,
4225-4239, 4226-4240, 4227-4241, 4228-4242, 4229-4243, 4230-4244,
4231-4245, 4232-4246, 4233-4247, 4234-4248, 4235-4249, 4236-4250,
4237-4251, 4238-4252, 4239-4253, 4240-4254, 4241-4255, 4242-4256,
4243-4257, 4244-4258, 4245-4259, 4246-4260, 4247-4261, 4248-4262,
4249-4263, 4250-4264, 4251-4265, 4252-4266, 4253-4267, 4254-4268,
4255-4269, 4256-4270, 4257-4271, 4258-4272, 4259-4273, 4260-4274,
4261-4275, 4262-4276, 4263-4277, 4264-4278, 4265-4279, 4266-4280,
4267-4281, 4268-4282, 4269-4283, 4270-4284, 4271-4285, 4272-4286,
4273-4287, 4274-4288, 4275-4289, 4276-4290, 4277-4291, 4278-4292,
4279-4293, 4280-4294, 4281-4295, 4282-4296, 4283-4297, 4284-4298,
4285-4299, 4286-4300, 4287-4301, 4288-4302, 4289-4303, 4290-4304,
4291-4305, 4292-4306, 4293-4307, 4294-4308, 4295-4309, 4296-4310,
4297-4311, 4298-4312, 4299-4313, 4300-4314, 4301-4315, 4302-4315,
4303-4317, 4304-4318, 4305-4319, 4306-4320, 4307-4321, 4308-4322,
4309-4323, 4310-4324, 4311-4325, 4312-4326, 4313-4327, 4314-4328,
4315-4329, 4316-4330, 4317-4331, 4318-4332, 4319-4333, 4320-4334,
4321-4335, 4322-4336, 4323-4337, 4324-4338, 4325-4339, 4326-4340,
4327-4341, 4328-4342, 4329-4343, 4330-4344, 4331-4345, 4332-4346,
4333-4347, 4334-4348, 4335-4349, 4336-4350, 4337-4351, 4338-4352,
4339-4353, 4340-4354, 4341-4355, 4342-4356, 4343-4357, 4344-4358,
4345-4359, 4346-4360, 4347-4361, 4348-4362, 4349-4363, 4350-4364,
4351-4365, 4352-4366, 4353-4367, 4354-4368, 4355-4369, 4356-4370,
4357-4371, 4358-4372, 4359-4373, 4360-4374, 4361-4375, 4362-4376,
4363-4377, 4364-4378, 4365-4379, 4366-4380, 4367-4381, 4368-4382,
4369-4383, 4370-4384, 4371-4385, 4372-4386, 4373-4387, 4374-4388,
4375-4389, 4376-4390, 4377-4391, 4378-4392, 4379-4393, 4380-4394,
4381-4395, 4382-4396, 4383-4397, 4384-4398, 4385-4399, 4386-4400,
4387-4401, 4388-4402, 4389-4403, 4390-4404, 4391-4405, 4392-4406,
4393-4407, 4394-4408, 4395-4409, 4396-4410, 4397-4411, 4398-4412,
4399-4413, 4400-4414, 4401-4415, 4402-4416, 4403-4417, 4404-4418,
4405-4419, 4406-4420, 4407-4421, 4408-4422, 4409-4423, 4410-4424,
4411-4425, 4412-4426, 4413-4427, 4414-4428, 4415-4429, 4416-4430,
4417-4431, 4418-4444, 4419-4433, 4420-4434, 4421-4435, 4422-4436,
4423-4437, 4424-4438, 4425-4439, 4426-4440, 4427-4441, 4428-4442,
4429-4443, 4430-4444, 4431-4445, 4432-4446, 4433-4447, 4434-4448,
4435-4449, 4436-4450, 4437-4451, 4438-4452, 4439-4453, 4440-4454,
4441-4455, 4442-4456, 4443-4457, 4444-4458, 4445-4459, 4446-4460,
4447-4461, 4448-4462, 4449-4463, 4450-4464, 4451-4465, 4452-4466,
4453-4467, 4454-4468, 4455-4469, 4456-4470, 4457-4471, 4458-4472,
4459-4473, 4460-4474, 4461-4475, 4462-4476, 4463-4477, 4464-4478,
4465-4479, 4466-4480, 4467-4481, 4468-4482, 4469-4483, 4470-4484,
4471-4485, 4472-4486, 4473-4487, 4474-4488, 4475-4489, 4476-4490,
4477-4491, 4478-4492, 4479-4493, 4480-4494, 4481-4495, 4482-4496,
4483-4497, 4484-4498, 4485-4499, 4486-4500, 4487-4501, 4488-4502,
4489-4503, 4490-4504, 4491-4505, 4492-4506, 4493-4507, 4494-4508,
4495-4509, 4496-4510, 4497-4511, 4498-4513, 4499-4514, 4500-4515,
4501-4515, 4502-4516, 4503-4517, 4504-4518, 4505-4519, 4506-4520,
4507-4521, 4508-4522, 4509-4523, 4510-4524, 4511-4525, 4512-4526,
4513-4527, 4514-4528, 4515-4529, 4516-4530, 4517-4531, 4518-4532,
4519-4533, 4520-4534, 4521-4535, 4522-4536, 4523-4537, 4524-4538,
4525-4539, 4526-4540, 4527-4541, 4528-4542, 4529-4543, 4530-4544,
4531-4545, 4532-4546, 4533-4547, 4534-4548, 4535-4549, 4536-4550,
4537-4551, 4538-4552, 4539-4553, 4540-4554, 4541-4555, 4542-4556,
4543-4557, 4544-4558, 4545-4559, 4546-4560, 4547-4561, 4548-4562,
4549-4563, 4550-4564, 4551-4565, 4552-4566, 4553-4567, 4554-4568,
4555-4569, 4556-4570, 4557-4571, 4558-4572, 4559-4573, 4560-4574,
4561-4575, 4562-4576, 4563-4577, 4564-4578, 4565-4579, 4566-4580,
4567-4581, 4568-4582, 4569-4583, 4570-4584, 4571-4585, 4572-4586,
4573-4587, 4574-4588, 4575-4589, 4576-4590, 4577-4591, 4578-4592,
4579-4593, 4580-4594, 4581-4595, 4582-4596, 4583-4597, 4584-4598,
4585-4599, 4586-4600, 4587-4601, 4588-4602, 4589-4603, 4590-4604,
4591-4605, 4592-4606, 4593-4607, 4594-4608, 4595-4609, 4596-4610,
4597-4611, 4598-4612, 4599-4613, 4600-4614, 4601-4615, 4602-4616,
4603-4617, 4604-4618, 4605-4619, 4606-4620, 4607-4621, 4608-4622,
4609-4623, 4610-4624, 4611-4625, 4612-4626, 4613-4627, 4614-4628,
4615-4629, 4616-4630, 4617-4631, 4618-4632, 4619-4633, 4620-4634,
4621-4635, 4622-4636, 4623-4637, 4624-4638, 4625-4639, 4626-4640,
4627-4641, 4628-4642, 4629-4643, 4630-4644, 4631-4645, 4632-4646,
4633-4647, 4634-4648, 4635-4649, 4636-4650, 4637-4651, 4638-4652,
4639-4653, 4640-4654, 4641-4655, 4642-4656, 4643-4657, 4644-4658,
4645-4659, 4646-4660, 4647-4661, 4648-4662, 4649-4663, 4650-4664,
4651-4665, 4652-4666, 4653-4667, 4654-4668, 4655-4669, 4656-4670,
4657-4671, 4658-4672, 4659-4673, 4660-4674, 4661-4675, 4662-4676,
4663-4677, 4664-4678, 4665-4679, 4666-4680, 4667-4681, 4668-4682,
4669-4683, 4670-4684, 4671-4685, 4672-4686, 4673-4687, 4674-4688,
4675-4689, 4676-4690, 4677-4691, 4678-4692, 4679-4693, 4680-4694,
4681-4695, 4682-4696, 4683-4697, 4684-4698, 4685-4699, 4686-4700,
4687-4701, 4688-4702, 4689-4703, 4690-4704, 4691-4705, 4692-4706,
4693-4707, 4694-4708, 4695-4709, 4696-4710, 4697-4711, 4698-4712,
4699-4713, 4700-4714, 4701-4715, 4702-4716, 4703-4717, 4704-4718,
4705-4719, 4706-4720, 4707-4721, 4708-4722, 4709-4723, 4710-4724,
4711-4725, 4712-4726, 4713-4727, 4714-4728, 4715-4729, 4716-4730,
4717-4731, 4718-4732, 4719-4733, 4720-4734, 4721-4735, 4722-4736,
4723-4737, 4724-4738, 4725-4739, 4726-4740, 4727-4741, 4728-4742,
4729-4743, 4730-4744, 4731-4745, 4732-4746, 4733-4747, 4734-4748,
4735-4749, 4736-4750, 4737-4751, 4738-4752, 4739-4753, 4740-4754,
4741-4755, 4742-4756, 4743-4757, 4744-4758, 4745-4759, 4746-4760,
4747-4761, 4748-4762, 4749-4763, 4750-4764, 4751-4765, 4752-4766,
4753-4767, 4754-4768, 4755-4769, 4756-4770, 4757-4771, 4758-4772,
4759-4773, 4760-4774, 4761-4775, 4762-4776, 4763-4777, 4764-4778,
4765-4779, 4766-4780, 4767-4781, 4768-4782, 4769-4783, 4770-4784,
4771-4785, 4772-4786, 4773-4787, 4774-4788, 4775-4789, 4776-4790,
4777-4791, 4778-4792, 4779-4793, 4780-4794, 4781-4795, 4782-4796,
4783-4797, 4784-4798, 4785-4799, 4786-4800, 4787-4801, 4788-4802,
4789-4803, 4790-4804, 4791-4805, 4792-4806, 4793-4807, 4794-4808,
4795-4809, 4796-4810, 4797-4811, 4798-4812, 4799-4813, 4800-4814,
4801-4815, 4802-4816, 4803-4817, 4804-4818, 4805-4819, 4806-4820,
4807-4821, 4808-4822, 4809-4823, 4810-4824, 4811-4825, 4812-4826,
4813-4827, 4814-4828, 4815-4829, 4816-4830, 4817-4831, 4818-4832,
4819-4833, 4820-4834, 4821-4835, 4822-4836, 4823-4837, 4824-4838,
4825-4839, 4826-4840, 4827-4841, 4828-4842, 4829-4843, 4830-4844,
4831-4845, 4832-4846, 4833-4847, 4834-4848, 4835-4849, 4836-4850,
4837-4851, 4838-4852, 4839-4853, 4840-4854, 4841-4855, 4842-4856,
4843-4857, 4844-4858, 4845-4859, 4846-4860, 4847-4861, 4848-4862,
4849-4863, 4850-4864, 4851-4865, 4852-4866, 4853-4867, 4854-4868,
4855-4869, 4856-4870, 4857-4871, 4858-4872, 4859-4873, 4860-4874,
4861-4875, 4862-4876, 4863-4877, 4864-4878, 4865-4879, 4866-4880,
4867-4881, 4868-4882, 4869-4883, 4870-4884, 4871-4885, 4872-4886,
4873-4887, 4874-4888, 4875-4889, 4876-4890, 4877-4891, 4878-4892,
4879-4893, 4880-4894, 4881-4895, 4882-4896, 4883-4897, 4884-4898,
4885-4899, 4886-4900, 4887-4901, 4888-4902, 4889-4903, 4890-4904,
4891-4905, 4892-4906, 4893-4907, 4894-4908, 4895-4909, 4896-4910,
4897-4911, 4898-4912, 4899-4913, 4900-4914, 4901-4915, 4902-4916,
4903-4917, 4904-4918, 4905-4919, 4906-4920, 4907-4921, 4908-4922,
4909-4923, 4910-4924, 4911-4925, 4912-4926, 4913-4927, 4914-4928,
4915-4929, 4916-4930, 4917-4931, 4918-4932, 4919-4933, 4920-4934,
4921-4935, 4922-4936, 4923-4937, 4924-4938, 4925-4939, 4926-4940,
4927-4941, 4928-4942, 4929-4943, 4930-4944, 4931-4945, 4932-4946,
4933-4947, 4934-4948, 4935-4949, 4936-4950, 4937-4951, 4938-4952,
4939-4953, 4940-4954, 4941-4955, 4942-4956, 4943-4957, 4944-4958,
4945-4959, 4946-4960, 4947-4961, 4948-4962, 4949-4963, 4950-4964,
4951-4965, 4952-4966, 4953-4967, 4954-4968, 4955-4969, 4956-4970,
4957-4971, 4958-4972, 4959-4973, 4960-4974, 4961-4975, 4962-4976,
4963-4977, 4964-4978, 4965-4979, 4966-4980, 4967-4981, 4968-4982,
4969-4983, 4970-4984, 4971-4985, 4972-4986, 4973-4987, 4974-4988
and 4975-4989 of SEQ ID NO:3.
EXAMPLE 9
INHIBITION OF IGF-I BINDING BY ANTISENSE OLIGONUCLEOTIDES TO IGF-I
RECEPTOR
Sub-confluent HaCaT cells were treated as described above with
phosphorothioate oligonucleotides IGFR.AS (antisense:
5'-ATCTCTCCGCTTCCTTTC-3'; [SEQ ID NO. 10]; ref 13) and IGFR.S
(sense control: 5'-GAAAGGAAGCGGAGAGAT-3'; [SEQ ID NO. 11]; ref 13)
IGF-I binding to the cell monolayers was then measured as .sup.125
I-IGF-I.
EXAMPLE 10
INHIBITION OF IGFBP-3 PRODUCTION USING ANTISENSE
OLIGONUCLEOTIDES
The results of this experiment are shown in FIGS. 7 and 8.
HaCaT cells were initially plated in DMEM with 10% v/v serum, then
AS oligo experiments were performed in complete "Keratinocyte-SFM"
(Gibco) to exclude the influence of exogenous IGFBPs. Oligos were
synthesised as phosphorothioate (nuclease-resistant) derivatives
(Bresatec, South Australia) and were as follows: antisense: AS2
(SEQ ID NO:4), 5'-GCGCCCGCTGCATGACGCCTGCAAC-3' (IGFBP-3 start
codon); controls: AS2NS (SEQ ID NO:8),
5'-CGGAGATGCCGCATGCCAGCGCAGG-3'; AS4 (SEQ ID NO:6),
5'-AGGCGGCTGACGGCACTA-3'; AS4NS (SEQ ID NO:9),
5'-GACAGCGTCGGAGCGATC-3'; IGFRAS (SEQ ID NO:10),
5'-ATCTCTCCGCTTCCTTTC-3'; IGFRS (SEQ ID NO:11),
5'-GAAAGGAAGCGGAGAGAT-3'. Oligos to IGFBP-3 were based on the
published sequence of Spratt et al [12]. AS oligos were added to
HaCaT monolayers in 0.5 ml medium in 24-well plates at the
concentrations and addition frequencies indicated. IGFBP-3 measured
in cell-conditioned medium using a dot-blot assay, adapted from the
Western ligand blot method of Hossenlopp et al [11], in which 100
.mu.l of conditioned medium was applied to nitrocellulose filters
with a vacuum dot-blot apparatus. After drying the membranes at
37.degree. C., relative amounts of IGFBP are determined by .sup.125
I-IGF-I-binding, autoradiography and computerised imaging
densitometry. Triplicate wells (except in FIG. 7, where duplicate
wells were measured as shown) were analysed and corrected for
changes in cell number per well. Relative cell number per well was
determined using an amido black dye method, developed specifically
for cultured monolayers of HaCaT cells (14). Cell numbers differed
by less than 10% after treatment. For oligos to the IGF receptor,
receptor quantitation in intact HaCaT monolayers was by overnight
incubation with .sup.125 I-IGF-I (30,000 cpm/well) at 4.degree.
C.
EXAMPLE 11
INHIBITION OF IGFBP-2 PRODUCTION USING RIBOZYMES
Experiments involving ribozymes are generally conducted as
described in International Patent Application No. WO 89/05852 and
in Haselhoff and Gerlach [8]. Ribozymes are constructed with a
hybridising region which is complementary in nucleotide sequence to
at least part of a target RNA which, in this case, encodes IGFBP-2.
Activity of ribozymes is measurable on, for example, Northern blots
or using animal models such as in the nude mouse model (15; 16) or
the "flaky skin" mouse model (17; 18).
EXAMPLE 12
INHIBITION OF IGFBP-3 PRODUCTION USING RIBOZYMES
The methods described in Example 11 are used for the screening of
ribozymes which inhibit IGFBP-3 production. The activity of the
ribozymes is determined as in Example 11.
EXAMPLE 13
INHIBITION OF IGF-1 PRODUCTION USING RIBOZYMES
The methods described in Example 11 are used for the screening of
ribozymes which inhibit IGF-1 production. The activity of the
ribozymes is determined as in Example 11.
EXAMPLE 14
INHIBITION OF IGF-1 RECEPTOR PRODUCTION USING RIBOZYMES
The methods described in Example 11 are used for the screening of
ribozymes which inhibit IGF-1 production. The activity of the
ribozymes is determined as in Example 11.
Those skilled in the art will appreciate at the invention described
herein is susceptible to variations and modifications other than
those specifically described. It is to be understood that the
invention includes all such variations and modifications. The
invention also includes all of the steps, features, compositions
and compounds referred to or indicated in this specification,
individually or collectively, and any and all combinations of any
two or more of said steps or features.
REFERENCES:
1. Sara V. Physiological Reviews 70: 591-614, 1990.
2. Rechler M. M. and Brown A. L. Growth Regulation 2: 55-68,
1992.
3. Clemmons D. R. Growth Regn 2: 80, 1992.
4. Oakes S. R., K. M. Haynes, M. J. Waters, A. C. Herington and G.
A. Werther J. Clin Endocrinol Metab 73: 1368-1373, 1992.
5. Camacho-Hubner C. et al. J. Biol Chem 267: 11949-11956,
1992.
6. Neely K. E. et al. J. Inv Derm 96: 104, 1991.
7. Ts'O POP, Aurelian L., Chang E. and Miller P. S. Nonionic
oligonucleotide analogs (Matagen TM) as anticodic agents in duplex
and triplex formation. in "Antisense Strategies", Annals of the New
York Academy of Sciences 660: 159-177 (Baserga R. and Denhardt D.
T., eds.), 1993.
8. Haseloff J. and Gerlach L. Nature 334: 586-591, 1988.
9. Boukamp P., Petrussevska R. T., Breitkreuz D., Hornung J.,
Markham A., Fusenig N. E. J. Cell Biol 106: 761-771, 1988.
10. Rheinwald and Green Cell 6: 331-344, 1975.
11. Hossenlopp P., Seurin D., Segovia-Quinson B., Hardouin S.,
Binoux M. Anal Biochem 154: 138-143, 1986.
12. Spratt S. K., Tatsuno G. P., Yamanaka M. K., Ark B. C., Detmer
J., Mascarenhas D., Flynn J., Talkington-Verser C., Spencer E. M.
Growth Factors 3: 63-72, 1990.
13. Pietrzkowski, Z., Sell C., Lammers R., Ullrich A. and Baserga
R. Mol. Cell. Biol. 12: 3883-3889, 1992.
14. Schulz J., Dettlaff S., Fritzsche U., Harms U., Schiebel H.,
Derer W., Fusenig N. E., Hulsen A. and Bohm M. J. Immunol. Meth.
167: 1-13, 1994.
15. Baker B. S., Brent L., Valdimarsson H., Powles A. V., Al-Imara
L., Walker M. and Fry L. Brit. J. Bermatol 126: 105-110, 1992.
16. Nanney L. B. et al J. Invest. Bermatol 98: 296-301, 1992.
17. Sundberg J. P. et al Immunol. Investigations 22: 389-401,
1993.
18. Sundberg J. P. et al J. Invest. Dermatol 102: 781-788,
1994.
__________________________________________________________________________
# SEQUENCE LISTING - (1) GENERAL INFORMATION: - (iii) NUMBER OF
SEQUENCES: 11 - (2) INFORMATION FOR SEQ ID NO:1: - (i) SEQUENCE
CHARACTERISTICS: #pairs (A) LENGTH: 1433 base (B) TYPE: nucleic
acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear - (ii) MOLECULE
TYPE: cDNA - (ix) FEATURE: (A) NAME/KEY: Coding Se - #quence (B)
LOCATION: 118...1101 (D) OTHER INFORMATION: (A) NAME/KEY: Coding Se
- #quence (B) LOCATION: 118...234 (D) OTHER INFORMATION: (A)
NAME/KEY: Coding Se - #quence (B) LOCATION: 235...1101 (D) OTHER
INFORMATION: - (x) PUBLICATION INFORMATION: (A) AUTHORS: Binkert, C
- #., et al. (B) TITLE: Cloning, seq - #uence analysis and
expression... (C) JOURNAL: EMBO J. (D) VOLUME: 8 (E) ISSUE: 1989
(F) PAGES: 2497-2502 (G) DATE: - (xi) SEQUENCE DESCRIPTION: SEQ ID
NO:1: #GGGCCGTGCA 60GAGGAGG AAGAAGCGGA GGAGGCGGCT CCCGCTCGCA
#CGCCAGC ATG 120CCGCTCG CTCGCTCGCC CGCCGCGCCG CGCTGCCGAC # Met # 1
#CTG CCG CCG CCG CCG CTG 168CG CTG CCG #Leu Pro Pro Pro Pro Leu Pro
Ala Leu Pro # 15 #CTG GGC GCG AGT GGC GGC 216TG CTG CTA #Leu Gly
Ala Ser Gly Gly Leu Leu Leu Leu # 30 #CGC TGC CCG CCC TGC ACA 264TG
CTG TTC #Arg Cys Pro Pro Cys Thr Glu Val Leu Phe # 45 #CCG GTT GCG
CCG CCC GCC 312GG CCC CCG #Pro Val Ala Pro Pro Ala Cys Gly Pro Pro
# 65 #ATG CCA TGC GCG GAG CTC 360GC GCC CGC #Met Pro Cys Ala Glu
Leu Gly Gly Ala Arg # 80 #GTG TGC GCC CGG CTG GAG 408GC TGC TCG
#Val Cys Ala Arg Leu Glu Gly Cys Cys Ser # 95 #TGC GGC CAG GGG CTG
CGC 456CC CCG CGC #Cys Gly Gln Gly Leu Arg Tyr Thr Pro Arg # 110
#CTG CAG GCG CTG GTC ATG 504AG CTG CCC #Leu Gln Ala Leu Val Met Ser
Glu Leu Pro # 125 #GCC GAG TAT GGC GCC AGC 552GC CGG GAC #Ala Glu
Tyr Gly Ala Ser Lys Arg Arg Asp #145 #CAC TCA GAA GGA GGC CTG 600GC
GAT GAC #His Ser Glu Gly Gly Leu Asn Gly Asp Asp # 160 #ATG TTG GGC
GGG GGA GGC 648CC ATG AAC #Met Leu Gly Gly Gly Gly Ser Thr Met Asn
# 175 #ATG AAG GAG CTG GCC GTG 696AG TCG GGT #Met Lys Glu Leu Ala
Val Leu Lys Ser Gly # 190 #CAG ATG GGC AAG GGT GGC 744AG CAC CGG
#Gln Met Gly Lys Gly Gly Glu Gln His Arg # 205 #AAG CTG CGA CCA CCC
CCT 792AG CCC AAG #Lys Leu Arg Pro Pro Pro Glu Glu Pro Lys #225
#CAG GTC CTG GAG CGG ATC 840AA CTG GAC #Gln Val Leu Glu Arg Ile Gln
Glu Leu Asp # 240 #CCT CTG GAG CAC CTC TAC 888AG CGG GGC #Pro Leu
Glu His Leu Tyr Asp Glu Arg Gly # 255 #GGC CTG TAC AAC CTC AAA
936AC AAG CAT #Gly Leu Tyr Asn Leu Lys Cys Asp Lys His # 270 #GGG
GAG TGC TGG TGT GTG 984GG CAG CGT #Gly Glu Cys Trp Cys Val Asn Gly
Gln Arg # 285 #GCC CCC ACC ATC CGG GGG 1032TC CAG GGA #Ala Pro Thr
Ile Arg Gly Leu Ile Gln Gly #305 #CAG CAG GAG GCT TGC GGG 1080AC
AAT GAG #Gln Gln Glu Ala Cys Gly Phe Tyr Asn Glu # 320 - GTG CAC
ACC CAG CGG ATG CAG TAGACCGCAG CCAGC - #CGGTG CCTGGCGCCC CTGC 1135
Val His Thr Gln Arg Met Gln 325 #GGGTGCTGGA 1195CCAAACA CCGGCAGAAA
ACGGAGAGTG CTTGGGTGGT #CGAGCTCGGC 1255CTGACAC ACGTATTTAT ATTTGGAAAG
AGACCAGCAC #TTGCTTTCCC 1315CTCTCTT CCCAGCTGCA GATGCCACAC CTGCTCCTTC
#GGGGGAAGAG 1375GGGTTGT GGTCGGGGAG CTGGGGTACA GGTTTGGGGA #GGAAAAGT
1433TGAACCC CTGTGTCCCT TTTGCATAAG ATTAAAGGAA - (2) INFORMATION FOR
SEQ ID NO:2: - (i) SEQUENCE CHARACTERISTICS: #pairs (A) LENGTH:
2474 base (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D)
TOPOLOGY: linear - (ii) MOLECULE TYPE: cDNA - (ix) FEATURE: (A)
NAME/KEY: Coding Se - #quence (B) LOCATION: 110...982 (D) OTHER
INFORMATION: - (x) PUBLICATION INFORMATION: (A) AUTHORS: Wood, W.I.
- #, et al. #expression of the growth... and (C) JOURNAL: Mol.
Endoc - #rinol. (D) VOLUME: 2 (E) ISSUE: 1988 (F) PAGES: 1176-1185
(G) DATE: - (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2: #CTGTCGCCCC
60CGCTTCC TGCCTGGATT CCACAGCTTC GCGCCGTGTA #ATG CAG CGG 118AGCCTG
CCAAGCAGCG TGCCCCGGTT GCAGGCGTC # Met Gln Arg # 1 #ACT CTG CTG GTG
CTG CTC 166CT GCG CTG #Thr Leu Leu Val Leu Leu Ala Ala Ala Leu # 15
#AGC TCG GGG GGC TTG GGT 214CT GGC GCG #Ser Ser Gly Gly Leu Gly Arg
Ala Gly Ala # 35 #CGT GCA CTG GCC CAG TGC 262GC GAC GCG #Arg Ala
Leu Ala Gln Cys Pro Cys Asp Ala # 50 #GTG CGC GAG CCG GGC TGC 310CG
GAG CTG #Val Arg Glu Pro Gly Cys Cys Ala Glu Leu # 65 #GGC CAG CCG
TGC GGC ATC 358TG AGC GAG #Gly Gln Pro Cys Gly Ile Ala Leu Ser Glu
# 80 #TGC CAG CCG TCG CCC GAC 406GC CTT CGC #Cys Gln Pro Ser Pro
Asp Ser Gly Leu Arg # 95 #GGC CGC GGG CTC TGC GTC 454TG CTG GAC
#Gly Arg Gly Leu Cys Val Ala Leu Leu Asp #115 #TAC CTG CTG CCA GCG
CCG 502TG CGC GCC #Tyr Leu Leu Pro Ala Pro Arg Leu Arg Ala # 130
#GAA GAC CGC AGC GCC GGC 550AG TCG GAG #Glu Asp Arg Ser Ala Gly Ser
Glu Ser Glu # 145 #CAC CGG GTG TCT GAT CCC 598CC AGC ACG #His Arg
Val Ser Asp Pro Val Ser Ser Thr # 160 #ATC ATC AAG AAA GGG CAT
646AG ATA ATC #Ile Ile Lys Lys Gly His Ser Lys Ile Ile # 175 #TAC
GAG TCT CAG AGC ACA 694AA GTT GAC #Tyr Glu Ser Gln Ser Thr Tyr Lys
Val Asp #195 #CGG GAG ACA GAA TAT GGT 742AG TCC AAG #Arg Glu Thr
Glu Tyr Gly Ser Glu Ser Lys # 210 #AAT CAC CTG AAG TTC CTC 790AC
ACA CTG #Asn His Leu Lys Phe Leu Glu Asp Thr Leu # 225 #CCC AAC TGT
GAC AAG AAG 838TA CAC ATT #Pro Asn Cys Asp Lys Lys Gly Val His Ile
# 240 #TCC AAA GGC AGG AAG CGG 886GT CGC CCT #Ser Lys Gly Arg Lys
Arg Gln Cys Arg Pro # 255 #CAG CCT CTC CCA GGC TAC 934AG TAT GGG
#Gln Pro Leu Pro Gly Tyr Asp Lys Tyr Gly #275 #TAC AGC ATG CAG AGC
AAG T 983TG CAC TGC #Tyr Ser Met Gln Ser Lys Asp Val His Cys # 290
#AAAAGACTGC 1043AAGTTAA TGTGGAGCTC AAATATGCCT TATTTTGCAC
#TGGTGAACTG 1103AGCAGCT GGCTACAGCC TCGATTTATA TTTCTGTTTG
#TCTTTGAATG 1163CAAAGTT TAGAAAGAGG TTTTTGAAAT GCCTATGGTT
#AATTTTCTTG 1223TCTTTTC ACTTTCCAGT AGTCAGCAAA GAGCAGTTTG
#TGCGCCCGTG 1283AAATATT CAGAGACTCG AGCACAGCAC CCAGACTTCA
#TAGGCGGCTG 1343ATGTTGG TCGAAGCGGC CGACCACTGA CTTTGTGACT
#ACAGGCTTTA 1403GAGAACA CGCTTCACCC CCACTCCCCG TACAGTGCGC
#TGCTCCTGGA 1463AACCTTT AAACCCCGGT CATCCGGACA TCCCAACGCA
#AACCAAGAAG 1523TGTGGTG TCATTTCTGA AACAAGGGCG TGGATCCCTC
#ATAAGGTGGA 1583TCAAGTG ACCTGTACTG CTTGGGGACT ATTGGAGAAA
#GGAACCTATA 1643AAAAAAT ATGTATCTAA GAATGTTCTA GGGCACTCTG
#CCCAGTCGAA 1703CGGGCCC TCCTCTTCAG GAATCTTCCT GAAGACATGG
#ACGGGAGAGT 1763TTTTGCT GCGGCCCCGT GGGGTAGGAG GGACAGAGAG
#ACTGCAGAAA 1823CAGAGGC ATCACAAGTA ATGGCACAAT TCTTCGGATG
#AGCTCTTTAA 1883GTTCAAC AACTCAAGAC GAAGCTTATT TCTGAGGATA
#TAAAAAAGAA 1943TTTTCAT CTCTCATCTT TTGTCCTCCT TAGCACAATG
#GGACACTGGG 2003ACAGGAA GGAGGAATGG CTTGCTGGGG AGCCCATCCA
#TTTTCTTTAT 2063TCACCCA TGTTTGTTGA ACTTAGAGTC ATTCTCATGC
#ACATAGCCCC 2123ATGCAGA GAAGATATGT TCTTGTTAAC ATTGTATACA
#CTATATGATA 2183TCTATAC TAGATAATCC TAGATGAAAT GTTAGAGATG
#CTCTCCCGGA 2243GACTGAG GAAAGGAGCT CACGCCCAGA GACTGGGCTG
#TGCTGCAGAC 2303AAGGTCT GGCAAAGTCA GGCTCAGGGA GACTCTGCCC
#TCAAGACATT 2363CACGCTG CATAGAGCTC TCCTTGAAAA CAGAGGGGTC
#GTATTTTTGA 2423AGCTTTT CTTTATTTTT TTAACTTTTT GGGGGGAAAA #T
2474CAATGTA TTTATAAATA GTAAATAAAG TTTTTACCAT - (2) INFORMATION FOR
SEQ ID NO:3: - (i) SEQUENCE CHARACTERISTICS: #pairs (A) LENGTH:
4989 base (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D)
TOPOLOGY: linear - (ii) MOLECULE TYPE: cDNA - (iii) HYPOTHETICAL:
NO - (iv) ANTI-SENSE: NO - (v) FRAGMENT TYPE: - (vi) ORIGINAL
SOURCE: - (x) PUBLICATION INFORMATION: (A) AUTHORS: Ullrich, A -
#., et al. #growth factor I receptor...-like (C) JOURNAL: EMBO J.
(D) VOLUME: 5 (E) ISSUE: 1986 (F) PAGES: 2503-2512 (G) DATE: - (xi)
SEQUENCE DESCRIPTION: SEQ ID NO:3: #GTCTGGCTCC 60TGAGAAA GGGAATTTCA
TCCCAAATAA AAGGAATGAA #GCTCTCGCTC 120CGACCTC GCTGTGGGGG CTCCTGTTTC
TCTCCGCCGC #CTATCAGCAG 180GAGAAAT CTGCGGGCCA GGCATCGACA TCCGCAACGA
#GCTCATCTCC 240AGAACTG CACGGTGATC GAGGGCTACC TCCACATCCT #CGAGTACTTG
300ACCGCAG CTACCGCTTC CCCAAGCTCA CGGTCATTAC #CCTCACGGTC 360TGGCTGG
CCTCGAGAGC CTCGGAGACC TCTTCCCCAA #GACCAATCTC 420AACTCTT CTACAACTAC
GCCCTGGTCA TCTTCGAGAT #GATTGAGAAA 480TTTACAA CCTGAGGAAC ATTACTCGGG
GGGCCATCAG #TGCGGTGTCC 540GTTACCT CTCCACTGTG GACTGGTCCC TGATCCTGGA
#TCCAGGGACC 600TGGGGAA TAAGCCCCCA AAGGAATGTG GGGACCTGTG #CTACCGCTGC
660CGATGTG TGAGAAGACC ACCATCAACA ATGAGTACAA #GGCGTGCACC 720GCTGCCA
GAAAATGTGC CCAAGCACGT GTGGGAAGCG #TGACAACGAC 780GCTGCCA CCCCGAGTGC
CTGGGCAGCT GCAGCGCGCC #TGCCTGCCCG 840CTTGCCG CCACTACTAC TATGCCGGTG
TCTGTGTGCC #CGCCAACATC 900GGTTTGA GGGCTGGCGC TGTGTGGACC GTGACTTCTG
#GTGCATGCAG 960GCAGCGA CTCCGAGGGG TTTGTGATCC ACGACGGCGA #CCCTTGTGAA
1020GCTTCAT CCGCAACGGC AGCCAGAGCA TGTACTGCAT #TTCTGTTACT
1080AGGTCTG TGAGGAAGAA AAGAAAACAA AGACCATTGA #TAACATCCGA
1140TCCAAGG ATGCACCATC TTCAAGGGCA ATTTGCTCAT #GGTGGTGACG
1200TTGCTTC AGAGCTGGAG AACTTCATGG GGCTCATCGA #AAAAAACCTT
1260TCCGCCA TTCTCATGCC TTGGTCTCCT TGTCCTTCCT #CCTCGACAAC
1320GAGAGGA GCAGCTAGAA GGGAATTACT CCTTCTACGT #AGCAGGGAAA
1380AACTGTG GGACTGGGAC CACCGCAACC TGACCATCAA #GGAGGAAGTG
1440TCAATCC CAAATTATGT GTTTCCGAAA TTTACCGCAT #CGGGGAGAGA
1500GGCGCCA AAGCAAAGGG GACATAAACA CCAGGAACAA
#GAATCGCATC 1560GTGACGT CCTGCATTTC ACCTCCACCA CCACGTCGAA
#CTTCACCGTT 1620ACCGGTA CCGGCCCCCT GACTACAGGG ATCTCATCAG
#TGCCTGCGGC 1680CACCCTT TAAGAATGTC ACAGAGTATG ATGGGCAGGA
#GGAGCCCGGC 1740ACATGGT GGACGTGGAC CTCCCGCCCA ACAAGGACGT
#GGCTGTGACC 1800GGCTGAA GCCCTGGACT CAGTACGCCG TTTACGTCAA
#GTACATTCGC 1860AGAACGA CCATATCCGT GGGGCCAAGA GTGAGATCTT
#CTCCTCTTCT 1920TTCCTTC CATTCCCTTG GACGTTCTTT CAGCATCGAA
#TTACTACATT 1980AGTGGAA CCCTCCCTCT CTGCCCAACG GCAACCTGAG
#CTGCTCCAAA 2040GGCAGCC TCAGGACGGC TACCTTTACC GGCACAATTA
#GGTCACAGAG 2100TCAGGAA GTATGCCGAC GGCACCATCG ACATTGAGGA
#CCCCAAAACT 2160AGGTGTG TGGTGGGGAG AAAGGGCCTT GCTGCGCCTG
#TGAGAATTTC 2220AGGCCGA GAAGGAGGAG GCTGAATACC GCAAAGTCTT
#CATGCAAGTG 2280TCTTCGT GCCCAGACCT GAAAGGAAGC GGAGAGATGT
#CTACAACATC 2340TGTCCAG CCGAAGCAGG AACACCACGG CCGCAGACAC
#GGATAACAAG 2400AGCTGGA GACAGAGTAC CCTTTCTTTG AGAGCAGAGT
#TATCCACAGC 2460TTTCTAA CCTTCGGCCT TTCACATTGT ACCGCATCGA
#TGCAAGGACT 2520CTGAGAA GCTGGGCTGC AGCGCCTCCA ACTTCGTCTT
#AAGGCCTGAA 2580GAGCAGA TGACATTCCT GGGCCAGTGA CCTGGGAGCC
#TCTAATGTAT 2640TAAAGTG GCCGGAACCT GAGAATCCCA ATGGATTGAT
#ACAGGAATAC 2700GATCACA AGTTGAGGAT CAGCGAGAAT GTGTGTCCAG
#AGCCCGGATT 2760GGGCCAA GCTAAACCGG CTAAACCCGG GGAACTACAC
#CTATGTCCAG 2820TCTCTGG GAATGGGTCG TGGACAGATC CTGTGTTCTT
#CGCTGTCCTG 2880ATGAAAA CTTCATCCAT CTGATCATCG CTCTGCCCGT
#AAATAACAGC 2940GGTTGGT GATTATGCTG TACGTCTTCC ATAGAAAGAG
#CGCTGCTGAT 3000GAGTGCT GTATGCCTCT GTGAACCCGG AGTACTTCAG
#CCGGGAACTT 3060ATGAGTG GGAGGTGGCT CGGGAGAAGA TCACCATGAG
#GAAAGATGAA 3120TTGGGAT GGTCTATGAA GGAGTTGCCA AGGGTGTGGT
#TGAGAGGATT 3180TGGCCAT TAAAACAGTG AACGAGGCCG CAAGCATGCG
#GGTGCGATTG 3240AAGCTTC TGTGATGAAG GAGTTCAATT GTCACCATGT
#GACACGGGGC 3300CCCAAGG CCAGCCAACA CTGGTCATCA TGGAACTGAT
#AGTCCTAGCA 3360ATCTCCG GTCTCTGAGG CCAGAAATGG AGAATAATCC
#CATGGCATAC 3420GCAAGAT GATTCAGATG GCCGGAGAGA TTGCAGACGG
#GGTAGCCGAA 3480AGTTCGT CCACAGAGAC CTTGCTGCCC GGAATTGCAT
#GACAGACTAT 3540AAATCGG AGATTTTGGT ATGACGCGAG ATATCTATGA
#GTCCCTCAAG 3600GCAAAGG GCTGCTGCCC GTGCGCTGGA TGTCTCCTGA
#CTGGGAGATC 3660CCACTTA CTCGGACGTC TGGTCCTTCG GGGTCGTCCT
#TCGCTTCGTC 3720AGCAGCC CTACCAGGGC TTGTCCAACG AGCAAGTCCT
#TGAACTGATG 3780TTCTGGA CAAGCCAGAC AACTGTCCTG ACATGCTGTT
#CATCAGCAGC 3840AGTATAA CCCCAAGATG AGGCCTTCCT TCCTGGAGAT
#CGAGGAGAAC 3900TGGAGCC TGGCTTCCGG GAGGTCTCCT TCTACTACAG
#CGTCCCCCTG 3960CGGAGGA GCTGGACCTG GAGCCAGAGA ACATGGAGAG
#ACACAAGGCC 4020CCTCGTC CTCCCTGCCA CTGCCCGACA GACACTCAGG
#GAGACAGCCT 4080GCCCTGG GGTGCTGGTC CTCCGCGCCA GCTTCGACGA
#CCAGTCTTCG 4140ACGGGGG CCGCAAGAAC GAGCGGGCCT TGCCGCTGCC
#CGCGCAGCGG 4200TGGATCC TGAATCTGTG CAAACAGTAA CGTGTGCGCA
#CTCAGTGGAT 4260AGAGAGT TTTAACAATC CATTCACAAG CCTCCTGTAC
#CACATTTGGG 4320CTTGCTG CCCGCGGGAG ACAGCTTCTC TGCAGTAAAA
#TTAACTGACA 4380CAATATG CAAGCAGCTT TTTATTCCCT GCCCAAACCC
#AGAACCAGTC 4440ACCTTAA TGACAACACT TAATAGCAAC AGAGCACTTG
#GCTTCATAAC 4500CCTGTCC TTCCCTGTTC TCCCTTTCTC TCTCCTCTCT
#AGGAAGTGGC 4560CCACAAG TCCAGCTGGG AAGCCCTTTT TATCAGTTTG
#GTCATTACAA 4620CCATCCA ACCACTGTAC ACACCCGCCT GACACCGTGG
#GACATGAAAT 4680GATGGAA ATTTTTACCT TTATCTTTCA CCTTTCTAGG
#AATTTTGCCA 4740TCGTTCA TCCAAGGCTG TTACCATTTT AACGCTGCCT
#GAGGCATGGG 4800TCTCCCT CATCGGCCCG GCGCTGATTC CTCGTGTCCG
#CCGTCCATCC 4860TGGTTGC TCCATTTGAG AGACACGCTG GCGACACACT
#TTCTGACTAG 4920GTGCTGC TCAAGGCCAC AGGCACACAG GTCTCATTGC
#GGGGAGAAGC 4980GGAACTG GACACAATAG GTCTTTCTCT CAGTGAAGGT # 4989 -
(2) INFORMATION FOR SEQ ID NO:4: - (i) SEQUENCE CHARACTERISTICS:
#pairs (A) LENGTH: 25 base (B) TYPE: nucleic acid (C) STRANDEDNESS:
single (D) TOPOLOGY: linear - (ii) MOLECULE TYPE: cDNA - (iii)
HYPOTHETICAL: NO - (iv) ANTI-SENSE: NO - (v) FRAGMENT TYPE: - (vi)
ORIGINAL SOURCE: - (xi) SEQUENCE DESCRIPTION: SEQ ID NO:4: # 25
GCCT GCAAC - (2) INFORMATION FOR SEQ ID NO:5: - (i) SEQUENCE
CHARACTERISTICS: #pairs (A) LENGTH: 24 base (B) TYPE: nucleic acid
(C) STRANDEDNESS: single (D) TOPOLOGY: linear - (ii) MOLECULE TYPE:
cDNA - (iii) HYPOTHETICAL: NO - (iv) ANTI-SENSE: NO - (v) FRAGMENT
TYPE: - (vi) ORIGINAL SOURCE: - (xi) SEQUENCE DESCRIPTION: SEQ ID
NO:5: # 24AGCT GGCG - (2) INFORMATION FOR SEQ ID NO:6: - (i)
SEQUENCE CHARACTERISTICS: #pairs (A) LENGTH: 18 base (B) TYPE:
nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear - (ii)
MOLECULE TYPE: cDNA - (iii) HYPOTHETICAL: NO - (iv) ANTI-SENSE: NO
- (v) FRAGMENT TYPE: - (vi) ORIGINAL SOURCE: - (xi) SEQUENCE
DESCRIPTION: SEQ ID NO:6: # 18 TA - (2) INFORMATION FOR SEQ ID
NO:7: - (i) SEQUENCE CHARACTERISTICS: #pairs (A) LENGTH: 19 base
(B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY:
linear - (ii) MOLECULE TYPE: cDNA - (iii) HYPOTHETICAL: NO - (iv)
ANTI-SENSE: NO - (v) FRAGMENT TYPE: - (vi) ORIGINAL SOURCE: - (xi)
SEQUENCE DESCRIPTION: SEQ ID NO:7: # 19 GGC - (2) INFORMATION FOR
SEQ ID NO:8: - (i) SEQUENCE CHARACTERISTICS: #pairs (A) LENGTH: 25
base (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY:
linear - (ii) MOLECULE TYPE: cDNA - (iii) HYPOTHETICAL: NO - (iv)
ANTI-SENSE: NO - (v) FRAGMENT TYPE: - (vi) ORIGINAL SOURCE: - (xi)
SEQUENCE DESCRIPTION: SEQ ID NO:8: # 25 CAGC GCAGG - (2)
INFORMATION FOR SEQ ID NO:9: - (i) SEQUENCE CHARACTERISTICS: #pairs
(A) LENGTH: 18 base (B) TYPE: nucleic acid (C) STRANDEDNESS: single
(D) TOPOLOGY: linear - (ii) MOLECULE TYPE: cDNA - (iii)
HYPOTHETICAL: NO - (iv) ANTI-SENSE: NO - (v) FRAGMENT TYPE: - (vi)
ORIGINAL SOURCE: - (xi) SEQUENCE DESCRIPTION: SEQ ID NO:9: # 18 TC
- (2) INFORMATION FOR SEQ ID NO:10: - (i) SEQUENCE CHARACTERISTICS:
#pairs (A) LENGTH: 18 base (B) TYPE: nucleic acid (C) STRANDEDNESS:
single (D) TOPOLOGY: linear - (ii) MOLECULE TYPE: cDNA - (iii)
HYPOTHETICAL: NO - (iv) ANTI-SENSE: NO - (v) FRAGMENT TYPE: - (vi)
ORIGINAL SOURCE: - (xi) SEQUENCE DESCRIPTION: SEQ ID NO:10: # 18 TC
- (2) INFORMATION FOR SEQ ID NO:11: - (i) SEQUENCE CHARACTERISTICS:
#pairs (A) LENGTH: 18 base (B) TYPE: nucleic acid (C) STRANDEDNESS:
single (D) TOPOLOGY: linear - (ii) MOLECULE TYPE: cDNA - (iii)
HYPOTHETICAL: NO - (iv) ANTI-SENSE: NO - (v) FRAGMENT TYPE: - (vi)
ORIGINAL SOURCE: - (xi) SEQUENCE DESCRIPTION: SEQ ID NO:11: # 18 AT
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